Use of affordable technology for the sensitive and specific diagnosis of onchocerciasis (river blindness)
Author: Guzmán Laparra, Gabriel Eduardo
Date: 2002-11-01
Location: Hillarpsalen, Retzius väg 8, Karolinska Institutet
Time: 9.30
Department: Mikrobiologiskt och Tumörbiologiskt Centrum (MTC) / Microbiology and Tumor Biology Center (MTC)
View/ Open:
thesis.pdf (2.016Mb)
Abstract
Onchocerciasis or river blindness is the most common infectious cause of
blindness. Even in the absence of blindness, the skin effects caused are
of social stigma. It is the goal of the World Health Organization to
eradicate filariasis (including onchocerciasis). Eradication of
onchocerciasis is to be undertaken by mass treatment with tile
microfilaricide ivermectin, combined with vector control. To date, the
gold standard for diagnosis of onchocerciasis is through the
parasitological identification of the organism, Onchocerca volvulus. As
ivermectin treatment provokes a decrease in the microfilarial load of the
individuals, the sensitivity of the assay for detection would need to be
enhanced. Before and even after eradication is achieved, sensitive,
specific, quick and affordable methods will be necessary for adequate
diagnostics and epidemiological assessment. A number of recombinant
proteins have been identified for use in diagnosis but the results have
been disappointing. Using cocktails of recombinant proteins somewhat
improved the sensitivity. However, production of recombinant material has
proved to be a problem and costs have not even been discussed. Since
onchocerciasis is a disease endemic in less welloff countries, the cost
of reliable diagnostic methods is not a trivial matter. Therefore, there
is definitely a need to re- address this problem of developing improved
methods for monitoring the eradication programme in the context of
sensitivity, specificity, availability and affordability.
Using manual ion exchange chromatography, we have isolated a fraction of
low molecular weight proteins of 0. volvulus (designated PakF), derived
from adult female worms. This fraction, used in a Dot Blot Assay (DBA ,
), proved to be highly sensitive and specific for the diagnosis of
onchocerciasis in different geographic regions. We have tested the
PakF-DBA in Guatemala (where no other filaria has been described), in
Ghana (where other filariases are prevalent) and in an infectious Disease
referral center in Sweden (a country not endemic for filaria).
Additionally, we have tested for specificity using well-defined filarial
sera from areas non-endemic for onchocerciasis. When used to analyze
samples from different endemic scenarios, the PakF-DBA reached high
levels of sensitivity (96%) and specificity (98%). Compared to methods
that use recombinant material, the PakF-DBA offered similar or even
better sensitivity and specificity values, but without the high costs
involved in the production of recombinant proteins.
The PakF-DBA was worked out so it could provide specific and sensitive
results and yet provide an affordable alternative to the use of more
expensive or sophisticated diagnostic methods. In this regard, the method
was simplified to use eluted finger pricked blood instead of serum as the
source of antibodies to the PakF mixture. Using this technique, a
positive correlation was found between the results obtained with the
"gold-standard" skin snip test and the PakF-DBA, suggesting that the
PakFDBA could substitute, or at least provide a more sensitive tool when
the parasitological examination is not possible, due to the reluctance of
the individuals to undergo this procedure, or the lack of qualified
personnel that performs the skin snip.
A closer look at the biochemical properties of this mixture of native
proteins revealed that PakF is stable at room temperature, which in some
endemic settings could reach 30-35 C. IgG4 antibody response from
infected individuals is infrequent and did not improve upon sensitivity,
which adds an attractive feature in terms of costs, as anti-IgG4 reagents
tend to be expensive. Its stability at relatively high working
temperatures makes PakF attractive under field conditions. In addition,
heat inactivation of sera or eluted blood to reduce the risk of labile
blood borne viruses does not affect the response to PakF. The relatively
simple production of PakF, together with its stability, makes the
PakF-DBA a serious contender in the quest for a reliable, yet affordable
method for the diagnosis of onchocerciasis in areas where costs arc an
important issue arid the disease threatens with recrudescence, even after
years of control.
The results of this work speak for the potential of this protein mixture,
PakF, as a contending diagnostic tool that, when put into the right
format, could be a valuable tool for the monitoring of onchocerciasis
control measures.
List of papers:
I. Guzman GE, Akuffo HO, Lavebratt C, Lujan R (1997). "Differential immune response to Onchocerca volvulus: IgG4 antibody responses differ in onchocerciasis patients from Guatemala and Ghana. " Acta Trop 63(1): 15-31
Pubmed
II. Lavebratt C, Ljungstrom I, Guzman G, Thors C, Eriksson T, Akuffo HO (1997). "Evaluation of serological assays for diagnosis of onchocercosis. " Scand J Infect Dis 29(1): 65-70
Pubmed
III. Guzman GE, Lavebratt C, Lujan R, Akuffo H (2002). "Diagnosis of onchocerciasis using highly specific and sensitive native proteins. " Scand J Infect Dis 34(8): 583-90
Pubmed
IV. Guzman GE, Awadzi K, Opoku N, Narayanan RB, Akuffo HO (2002). "Comparison between the skin snip test and simple dot blot assay as potential rapid assessment tools for Onchocerciasis in the postcontrol era in Ghana. " Clin Diagn Lab Immunol 9(5): 1014-20
Pubmed
V. Guzman GE, Akuffo HO (2002). "Continued symptoms of onchocerciaasis 20 years after leaving the endemic area." (Submitted)
VI. Guzman GE, Akuffo HO (2002). "Biochemical characterization of the immunodominant proteins of PakF, a highly specific and sensitive mixture of native proteins used in the diagnosis of onchocerciasis." (Submitted)
I. Guzman GE, Akuffo HO, Lavebratt C, Lujan R (1997). "Differential immune response to Onchocerca volvulus: IgG4 antibody responses differ in onchocerciasis patients from Guatemala and Ghana. " Acta Trop 63(1): 15-31
Pubmed
II. Lavebratt C, Ljungstrom I, Guzman G, Thors C, Eriksson T, Akuffo HO (1997). "Evaluation of serological assays for diagnosis of onchocercosis. " Scand J Infect Dis 29(1): 65-70
Pubmed
III. Guzman GE, Lavebratt C, Lujan R, Akuffo H (2002). "Diagnosis of onchocerciasis using highly specific and sensitive native proteins. " Scand J Infect Dis 34(8): 583-90
Pubmed
IV. Guzman GE, Awadzi K, Opoku N, Narayanan RB, Akuffo HO (2002). "Comparison between the skin snip test and simple dot blot assay as potential rapid assessment tools for Onchocerciasis in the postcontrol era in Ghana. " Clin Diagn Lab Immunol 9(5): 1014-20
Pubmed
V. Guzman GE, Akuffo HO (2002). "Continued symptoms of onchocerciaasis 20 years after leaving the endemic area." (Submitted)
VI. Guzman GE, Akuffo HO (2002). "Biochemical characterization of the immunodominant proteins of PakF, a highly specific and sensitive mixture of native proteins used in the diagnosis of onchocerciasis." (Submitted)
Issue date: 2002-10-11
Rights:
Publication year: 2002
ISBN: 91-7349-321-x
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