An outbreak of acute haemorrhagic conjunctivitis occurred in The Gambia, West Africa in 2011. Aff... more An outbreak of acute haemorrhagic conjunctivitis occurred in The Gambia, West Africa in 2011. Affected individuals presented with conjunctival haemorrhages, swelling and ocular discharge. In an effort to identify a causative agent of the disease, ocular swabs were taken from patients during the acute and convalescent phases. Total RNA was extracted from all samples and reverse-transcriptase PCR performed using primers specific for all enteroviruses. Resulting amplicons were sequenced and data compared to known sequences using the BLAST algorithm. Forty-eight swabs were included in the analysis. Of these, 21 acute and 9 convalescent swabs (65% of the total) gave positive PCR results. Sequence analysis of the resulting amplicons indicated 99% sequence identity with coxsackievirus A24 variant identified during independent outbreaks of acute haemorrhagic conjunctivitis around the world and suggest the Gambian outbreak was due to this virus.
Trachoma, the leading infectious cause of blindness worldwide, is caused by conjunctival Chlamydi... more Trachoma, the leading infectious cause of blindness worldwide, is caused by conjunctival Chlamydia trachomatis infection. Trachoma is diagnosed clinically by observation of conjunctival inflammation and/or scarring; however, there is evidence that monitoring C. trachomatis infection may be required for elimination programmes. There are many commercial and 'in-house' nucleic acid amplification tests for the detection of C. trachomatis DNA, but the majority have not been validated for use with ocular swabs. This study evaluated a commercial assay, the Fast-Track Vaginal swab kit, using conjunctival samples from trachoma-endemic areas. An objective, biostatistical-based method for binary classification of continuous PCR data was developed, to limit potential user-bias in diagnostic settings. The Fast-Track Vaginal swab assay was run on 210 ocular swab samples from Guinea-Bissau and Tanzania. Fit of individual amplification curves to exponential or sigmoid models, derivative and...
Clinical signs of active (inflammatory) trachoma are found in many children in the Solomon Island... more Clinical signs of active (inflammatory) trachoma are found in many children in the Solomon Islands, but the majority of these individuals have no serological evidence of previous infection with Chlamydia trachomatis. In Temotu and Rennell and Bellona provinces, ocular infections with C. trachomatis were seldom detected among children with active trachoma; a similar lack of association was seen between active trachoma and other common bacterial and viral causes of follicular conjunctivitis. Here, we set out to characterise patterns of gene expression at the conjunctivae of children in these provinces with and without clinical signs of trachomatous inflammation-follicular (TF) and C. trachomatis infection. Purified RNA from children with and without active trachoma was run on Affymetrix GeneChip Human Transcriptome Array 2.0 microarrays. Profiles were compared between individuals with ocular C. trachomatis infection and TF (group DI; nā=ā6), individuals with TF but no C. trachomatis i...
Trachoma, a neglected tropical disease, is caused by ocular infection with Chlamydia trachomatis ... more Trachoma, a neglected tropical disease, is caused by ocular infection with Chlamydia trachomatis (Ct). The World Health Organization (WHO) recommends three annual rounds of community mass drug treatment with azithromycin (MDA) if the prevalence of follicular trachoma in 1-9 year olds (TF1-9) exceeds 10% at district level to achieve an elimination target of district-level TF1-9 below 5% after. To evaluate this strategy in treatment-naĆÆve trachoma-endemic island communities in Guinea Bissau, we conducted a cross-sectional population-based trachoma survey on four islands. The upper tarsal conjunctivae of each participant were clinically assessed for trachoma and conjunctival swabs were obtained (n = 1507). We used a droplet digital PCR assay to detect Ct infection and estimate bacterial load. We visited the same households during a second cross-sectional survey and repeated the ocular examination and obtained conjunctival swabs from these households one year after MDA (n = 1029). Pre-M...
Background: Several non-chlamydial microbial pathogens are associated with clinical signs of acti... more Background: Several non-chlamydial microbial pathogens are associated with clinical signs of active trachoma in trachoma endemic communities with a low prevalence of ocular Chlamydia trachomatis (Ct) infection. We observed a low prevalence of Ct in the Solomon Islands, therefore hypothesised that the high prevalence of active trachoma could be explained by a common non-chlamydial infection or by a dominant polymicrobial community dysbiosis. Methods: We studied DNA from conjunctival swabs collected from 257 Solomon Islanders with active trachoma and 257 matched controls. Droplet digital PCR was used to test for Adenoviridae, Haemophilus influenzae, Moraxella catarrhalis, Staphylococcus aureus, coagulase-negative Staphylococcus and Streptococcus pneumoniae. Polymicrobial community diversity and composition were studied by sequencing of hypervariable regions of the 16S ribosomal ribonucleic acid gene in a subset of 54 cases and 53 controls. Results: Although Ct was associated with acti...
Ocular Chlamydia trachomatis (Ct) infection causes trachoma, the leading infectious cause of blin... more Ocular Chlamydia trachomatis (Ct) infection causes trachoma, the leading infectious cause of blindness. A Ct D/UW3 proteome microarray and sera from Gambian adults with trachomatous trichiasis (TT) or healthy matched controls previously identified several novel antigens, which suggested differential recognition in adults with TT. We re-analysed this serological microarray data using more robust microarray analysis techniques accounting for typical problems associated with highly dimensional data. We examined the Ct-specific antibody profile concerning the overall diversity of responses, antigen expression stage and cellular localisation of antigens. We tested differentially recognised antigens by further serological testing of the screened sera and used larger independent sample sets for validation. Antibody responses identified High-Performance on antigens expressed early and late in the Ct developmental cycle and those secreted or localised to the outer membrane. Eight antigens we...
Chlamydia trachomatis (Ct) is the most common cause of bacterial sexually transmitted infection a... more Chlamydia trachomatis (Ct) is the most common cause of bacterial sexually transmitted infection and infectious cause of blindness (trachoma) worldwide. Understanding the spatial distribution of Ct infection may enable us to identify populations at risk and improve our understanding of Ct transmission. In this study, we sought to investigate the spatial distribution of Ct infection and the clinical features associated with high Ct load in trachoma-endemic communities on the BijagĆ³s Archipelago (Guinea Bissau). We collected 1507 conjunctival samples and corresponding detailed clinical data during a cross-sectional population-based geospatially representative trachoma survey. We used droplet digital PCR to estimate Ct load on conjunctival swabs. Geostatistical tools were used to investigate clustering of ocular Ct infections. Spatial clusters (independent of age and gender) of individuals with high Ct loads were identified using local indicators of spatial association. We did not detec...
Trachoma, caused by ocular infection with Chlamydia trachomatis, is the leading infectious cause ... more Trachoma, caused by ocular infection with Chlamydia trachomatis, is the leading infectious cause of blindness worldwide. We conducted the first population-based trachoma prevalence survey in the Casamance region of Senegal to enable the Senegalese National Eye Care Programme (NECP) to plan its trachoma control activities. The World Health Organization (WHO) guidelines state that any individual with trachomatous trichiasis (TT) should be offered surgery, but that surgery should be prioritised where the prevalence is >0.1%, and that districts and communities with a trachomatous inflammation, follicular (TF) prevalence of ā„10% in 1-9Ā year-olds should receive mass antibiotic treatment annually for a minimum of three years, along with hygiene promotion and environmental improvement, before re-assessing the prevalence to determine whether treatment can be discontinued (when TF prevalence in 1-9Ā year-olds falls <5%). Local healthcare workers conducted a population-based household sur...
Trachoma, caused by the intracellular bacterium Chlamydia trachomatis (Ct), is the leading infect... more Trachoma, caused by the intracellular bacterium Chlamydia trachomatis (Ct), is the leading infectious cause of preventable blindness. Many commercial platforms are available that provide highly sensitive and specific detection of Ct DNA. However, the majority of these commercial platforms are inaccessible for population-level surveys in resource-limited settings typical to trachoma control programmes. We developed two low-cost quantitative PCR (qPCR) tests for Ct using readily available reagents on standard real-time thermocyclers. Each multiplex qPCR test targets one genomic and one plasmid Ct target in addition to an endogenous positive control for Homo sapiens DNA. The quantitative performance of the qPCR assays in clinical samples was determined by comparison to a previously evaluated droplet digital PCR (ddPCR) test. The diagnostic performance of the qPCR assays were evaluated against a commercial assay (artus C. trachomatis Plus RG PCR, Qiagen) using molecular diagnostics qual...
Mass drug administration (MDA) with azithromycin is a corner-stone of trachoma control however it... more Mass drug administration (MDA) with azithromycin is a corner-stone of trachoma control however it may drive the emergence of antimicrobial resistance. In a cluster-randomized trial (Clinical trial gov NCT00792922), we compared the reduction in the prevalence of active trachoma in communities that received three annual rounds of MDA to that in communities that received a single treatment round. We used the framework of this trial to carry out an opportunistic study to investigate if the increased rounds of treatment resulted in increased prevalence of nasopharyngeal carriage of macrolide-resistant Staphylococcus aureus. Three cross-sectional surveys were conducted in two villages receiving three annual rounds of MDA (3āĆātreatment arm). Surveys were conducted immediately before the third round of MDA (CSS-1) and at one (CSS-2) and six (CSS-3) months after MDA. The final survey also included six villages that had received only one round of MDA 30Ā months previously (1āĆātreatment arm)....
We previously showed that conjunctival miR-147b and miR-1285 were upregulated in Gambian adults w... more We previously showed that conjunctival miR-147b and miR-1285 were upregulated in Gambian adults with inflammatory scarring trachoma, and miR-155 and miR-184 expression was strongly associated with conjunctival inflammation and ocular Chlamydia trachomatis infection in children from Guinea-Bissau. We investigated whether the single or combined expression of miR-147b, miR-1285, miR-155 and miR-184 was able to identify individuals with increased risk of incident or progressive scarring trachoma. Conjunctival swab samples were collected from 506 children between the ages of 4 and 12 living in northern Tanzania. These 506 samples formed the baseline sample set of a 4-year longitudinal study. ChlamydiaĀ trachomatis infection was diagnosed by droplet digital PCR and expression of miR-155, miR-184, miR-1285 and miR-147b was tested by qPCR. Individuals were assessed for incidence and progression of conjunctival scarring by comparison of conjunctival photographs taken at baseline and 4 years l...
Efforts are underway to eliminate trachoma as a public health problem by 2020. Programmatic guide... more Efforts are underway to eliminate trachoma as a public health problem by 2020. Programmatic guidelines are based on clinical signs that correlate poorly with Chlamydia trachomatis (Ct) infection in post-treatment and low-endemicity settings. Age-specific seroprevalence of anti Ct Pgp3 antibodies has been proposed as an alternative indicator of the need for intervention. To standardise the use of these tools, it is necessary to develop an analytical approach that performs reproducibly both within and between studies. Dried blood spots were collected in 2014 from children aged 1-9 years in Laos (n = 952) and Uganda (n = 2700) and from people aged 1-90 years in The Gambia (n = 1868). Anti-Pgp3 antibodies were detected by ELISA. A number of visual and statistical analytical approaches for defining serological status were compared. Seroprevalence was estimated at 11.3% (Laos), 13.4% (Uganda) and 29.3% (The Gambia) by visual inspection of the inflection point. The expectation-maximisation...
An outbreak of acute haemorrhagic conjunctivitis occurred in The Gambia, West Africa in 2011. Aff... more An outbreak of acute haemorrhagic conjunctivitis occurred in The Gambia, West Africa in 2011. Affected individuals presented with conjunctival haemorrhages, swelling and ocular discharge. In an effort to identify a causative agent of the disease, ocular swabs were taken from patients during the acute and convalescent phases. Total RNA was extracted from all samples and reverse-transcriptase PCR performed using primers specific for all enteroviruses. Resulting amplicons were sequenced and data compared to known sequences using the BLAST algorithm. Forty-eight swabs were included in the analysis. Of these, 21 acute and 9 convalescent swabs (65% of the total) gave positive PCR results. Sequence analysis of the resulting amplicons indicated 99% sequence identity with coxsackievirus A24 variant identified during independent outbreaks of acute haemorrhagic conjunctivitis around the world and suggest the Gambian outbreak was due to this virus.
Trachoma, the leading infectious cause of blindness worldwide, is caused by conjunctival Chlamydi... more Trachoma, the leading infectious cause of blindness worldwide, is caused by conjunctival Chlamydia trachomatis infection. Trachoma is diagnosed clinically by observation of conjunctival inflammation and/or scarring; however, there is evidence that monitoring C. trachomatis infection may be required for elimination programmes. There are many commercial and 'in-house' nucleic acid amplification tests for the detection of C. trachomatis DNA, but the majority have not been validated for use with ocular swabs. This study evaluated a commercial assay, the Fast-Track Vaginal swab kit, using conjunctival samples from trachoma-endemic areas. An objective, biostatistical-based method for binary classification of continuous PCR data was developed, to limit potential user-bias in diagnostic settings. The Fast-Track Vaginal swab assay was run on 210 ocular swab samples from Guinea-Bissau and Tanzania. Fit of individual amplification curves to exponential or sigmoid models, derivative and...
Clinical signs of active (inflammatory) trachoma are found in many children in the Solomon Island... more Clinical signs of active (inflammatory) trachoma are found in many children in the Solomon Islands, but the majority of these individuals have no serological evidence of previous infection with Chlamydia trachomatis. In Temotu and Rennell and Bellona provinces, ocular infections with C. trachomatis were seldom detected among children with active trachoma; a similar lack of association was seen between active trachoma and other common bacterial and viral causes of follicular conjunctivitis. Here, we set out to characterise patterns of gene expression at the conjunctivae of children in these provinces with and without clinical signs of trachomatous inflammation-follicular (TF) and C. trachomatis infection. Purified RNA from children with and without active trachoma was run on Affymetrix GeneChip Human Transcriptome Array 2.0 microarrays. Profiles were compared between individuals with ocular C. trachomatis infection and TF (group DI; nā=ā6), individuals with TF but no C. trachomatis i...
Trachoma, a neglected tropical disease, is caused by ocular infection with Chlamydia trachomatis ... more Trachoma, a neglected tropical disease, is caused by ocular infection with Chlamydia trachomatis (Ct). The World Health Organization (WHO) recommends three annual rounds of community mass drug treatment with azithromycin (MDA) if the prevalence of follicular trachoma in 1-9 year olds (TF1-9) exceeds 10% at district level to achieve an elimination target of district-level TF1-9 below 5% after. To evaluate this strategy in treatment-naĆÆve trachoma-endemic island communities in Guinea Bissau, we conducted a cross-sectional population-based trachoma survey on four islands. The upper tarsal conjunctivae of each participant were clinically assessed for trachoma and conjunctival swabs were obtained (n = 1507). We used a droplet digital PCR assay to detect Ct infection and estimate bacterial load. We visited the same households during a second cross-sectional survey and repeated the ocular examination and obtained conjunctival swabs from these households one year after MDA (n = 1029). Pre-M...
Background: Several non-chlamydial microbial pathogens are associated with clinical signs of acti... more Background: Several non-chlamydial microbial pathogens are associated with clinical signs of active trachoma in trachoma endemic communities with a low prevalence of ocular Chlamydia trachomatis (Ct) infection. We observed a low prevalence of Ct in the Solomon Islands, therefore hypothesised that the high prevalence of active trachoma could be explained by a common non-chlamydial infection or by a dominant polymicrobial community dysbiosis. Methods: We studied DNA from conjunctival swabs collected from 257 Solomon Islanders with active trachoma and 257 matched controls. Droplet digital PCR was used to test for Adenoviridae, Haemophilus influenzae, Moraxella catarrhalis, Staphylococcus aureus, coagulase-negative Staphylococcus and Streptococcus pneumoniae. Polymicrobial community diversity and composition were studied by sequencing of hypervariable regions of the 16S ribosomal ribonucleic acid gene in a subset of 54 cases and 53 controls. Results: Although Ct was associated with acti...
Ocular Chlamydia trachomatis (Ct) infection causes trachoma, the leading infectious cause of blin... more Ocular Chlamydia trachomatis (Ct) infection causes trachoma, the leading infectious cause of blindness. A Ct D/UW3 proteome microarray and sera from Gambian adults with trachomatous trichiasis (TT) or healthy matched controls previously identified several novel antigens, which suggested differential recognition in adults with TT. We re-analysed this serological microarray data using more robust microarray analysis techniques accounting for typical problems associated with highly dimensional data. We examined the Ct-specific antibody profile concerning the overall diversity of responses, antigen expression stage and cellular localisation of antigens. We tested differentially recognised antigens by further serological testing of the screened sera and used larger independent sample sets for validation. Antibody responses identified High-Performance on antigens expressed early and late in the Ct developmental cycle and those secreted or localised to the outer membrane. Eight antigens we...
Chlamydia trachomatis (Ct) is the most common cause of bacterial sexually transmitted infection a... more Chlamydia trachomatis (Ct) is the most common cause of bacterial sexually transmitted infection and infectious cause of blindness (trachoma) worldwide. Understanding the spatial distribution of Ct infection may enable us to identify populations at risk and improve our understanding of Ct transmission. In this study, we sought to investigate the spatial distribution of Ct infection and the clinical features associated with high Ct load in trachoma-endemic communities on the BijagĆ³s Archipelago (Guinea Bissau). We collected 1507 conjunctival samples and corresponding detailed clinical data during a cross-sectional population-based geospatially representative trachoma survey. We used droplet digital PCR to estimate Ct load on conjunctival swabs. Geostatistical tools were used to investigate clustering of ocular Ct infections. Spatial clusters (independent of age and gender) of individuals with high Ct loads were identified using local indicators of spatial association. We did not detec...
Trachoma, caused by ocular infection with Chlamydia trachomatis, is the leading infectious cause ... more Trachoma, caused by ocular infection with Chlamydia trachomatis, is the leading infectious cause of blindness worldwide. We conducted the first population-based trachoma prevalence survey in the Casamance region of Senegal to enable the Senegalese National Eye Care Programme (NECP) to plan its trachoma control activities. The World Health Organization (WHO) guidelines state that any individual with trachomatous trichiasis (TT) should be offered surgery, but that surgery should be prioritised where the prevalence is >0.1%, and that districts and communities with a trachomatous inflammation, follicular (TF) prevalence of ā„10% in 1-9Ā year-olds should receive mass antibiotic treatment annually for a minimum of three years, along with hygiene promotion and environmental improvement, before re-assessing the prevalence to determine whether treatment can be discontinued (when TF prevalence in 1-9Ā year-olds falls <5%). Local healthcare workers conducted a population-based household sur...
Trachoma, caused by the intracellular bacterium Chlamydia trachomatis (Ct), is the leading infect... more Trachoma, caused by the intracellular bacterium Chlamydia trachomatis (Ct), is the leading infectious cause of preventable blindness. Many commercial platforms are available that provide highly sensitive and specific detection of Ct DNA. However, the majority of these commercial platforms are inaccessible for population-level surveys in resource-limited settings typical to trachoma control programmes. We developed two low-cost quantitative PCR (qPCR) tests for Ct using readily available reagents on standard real-time thermocyclers. Each multiplex qPCR test targets one genomic and one plasmid Ct target in addition to an endogenous positive control for Homo sapiens DNA. The quantitative performance of the qPCR assays in clinical samples was determined by comparison to a previously evaluated droplet digital PCR (ddPCR) test. The diagnostic performance of the qPCR assays were evaluated against a commercial assay (artus C. trachomatis Plus RG PCR, Qiagen) using molecular diagnostics qual...
Mass drug administration (MDA) with azithromycin is a corner-stone of trachoma control however it... more Mass drug administration (MDA) with azithromycin is a corner-stone of trachoma control however it may drive the emergence of antimicrobial resistance. In a cluster-randomized trial (Clinical trial gov NCT00792922), we compared the reduction in the prevalence of active trachoma in communities that received three annual rounds of MDA to that in communities that received a single treatment round. We used the framework of this trial to carry out an opportunistic study to investigate if the increased rounds of treatment resulted in increased prevalence of nasopharyngeal carriage of macrolide-resistant Staphylococcus aureus. Three cross-sectional surveys were conducted in two villages receiving three annual rounds of MDA (3āĆātreatment arm). Surveys were conducted immediately before the third round of MDA (CSS-1) and at one (CSS-2) and six (CSS-3) months after MDA. The final survey also included six villages that had received only one round of MDA 30Ā months previously (1āĆātreatment arm)....
We previously showed that conjunctival miR-147b and miR-1285 were upregulated in Gambian adults w... more We previously showed that conjunctival miR-147b and miR-1285 were upregulated in Gambian adults with inflammatory scarring trachoma, and miR-155 and miR-184 expression was strongly associated with conjunctival inflammation and ocular Chlamydia trachomatis infection in children from Guinea-Bissau. We investigated whether the single or combined expression of miR-147b, miR-1285, miR-155 and miR-184 was able to identify individuals with increased risk of incident or progressive scarring trachoma. Conjunctival swab samples were collected from 506 children between the ages of 4 and 12 living in northern Tanzania. These 506 samples formed the baseline sample set of a 4-year longitudinal study. ChlamydiaĀ trachomatis infection was diagnosed by droplet digital PCR and expression of miR-155, miR-184, miR-1285 and miR-147b was tested by qPCR. Individuals were assessed for incidence and progression of conjunctival scarring by comparison of conjunctival photographs taken at baseline and 4 years l...
Efforts are underway to eliminate trachoma as a public health problem by 2020. Programmatic guide... more Efforts are underway to eliminate trachoma as a public health problem by 2020. Programmatic guidelines are based on clinical signs that correlate poorly with Chlamydia trachomatis (Ct) infection in post-treatment and low-endemicity settings. Age-specific seroprevalence of anti Ct Pgp3 antibodies has been proposed as an alternative indicator of the need for intervention. To standardise the use of these tools, it is necessary to develop an analytical approach that performs reproducibly both within and between studies. Dried blood spots were collected in 2014 from children aged 1-9 years in Laos (n = 952) and Uganda (n = 2700) and from people aged 1-90 years in The Gambia (n = 1868). Anti-Pgp3 antibodies were detected by ELISA. A number of visual and statistical analytical approaches for defining serological status were compared. Seroprevalence was estimated at 11.3% (Laos), 13.4% (Uganda) and 29.3% (The Gambia) by visual inspection of the inflection point. The expectation-maximisation...
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