Application of antibodies to recombinant heat shock protein 70 in immunohistochemical diagnosis of mycobacterium avium subspecies paratuberculosis in tissues of naturally infected cattle

Table S1. Information regarding the samples included in the study. (DOCX 17 kb

Evaluation of Capilia TB assay for rapid identification of Mycobacterium tuberculosis complex in BACTEC MGIT 960 and BACTEC 9120 blood cultures

<p>Abstract</p> <p>Background</p> <p>Capilia TB is a simple immunochromatographic assay based on the detection of MPB64 antigen specifically secreted by the <it>Mycobacterium tuberculosis </it>complex (MTC). Capilia TB was evaluated for rapid identification of MTC from BACTEC MGIT 960 and BACTEC 9120 systems in Kampala, Uganda. Since most studies have mainly dealt with respiratory samples, the performance of Capilia TB on blood culture samples was also evaluated.</p> <p>Methods</p> <p>One thousand samples from pulmonary and disseminated tuberculosis (TB) suspects admitted to the JCRC clinic and the TB wards at Old Mulago hospital in Kampala, Uganda, were cultured in automated BACTEC MGIT 960 and BACTEC 9120 blood culture systems. BACTEC-positive samples were screened for purity by sub-culturing on blood agar plates. Two hundred and fifty three (253) samples with Acid fast bacilli (AFB, 174 BACTEC MGIT 960 and 79 BACTEC 9120 blood cultures) were analyzed for presence of MTC using Capilia TB and in-house PCR assays.</p> <p>Results</p> <p>The overall Sensitivity, Specificity, Positive and Negative Predictive values, and Kappa statistic for Capilia TB assay for identification of MTC were 98.4%, 97.6%, 97.7%, 98.4% and 0.96, respectively. Initially, the performance of in-house PCR on BACTEC 9120 blood cultures was poor (Sensitivity, Specificity, PPV, NPV and Kappa statistic of 100%, 29.3%,7%, 100% and 0.04, respectively) but improved upon sub-culturing on solid medium (Middlebrook 7H10) to 100%, 95.6%, 98.2%, 100% and 0.98, respectively. In contrast, the Sensitivity and Specificity of Capilia TB assay was 98.4% and 97.9%, respectively, both with BACTEC blood cultures and Middlebrook 7H10 cultured samples, revealing that Capilia was better than in-house PCR for identification of MTC in blood cultures. Additionally, Capilia TB was cheaper than in-house PCR for individual samples ($2.03 vs.$12.59, respectively), and was easier to perform with a shorter turnaround time (20 min vs. 480 min, respectively).</p> <p>Conclusion</p> <p>Capilia TB assay is faster and cheaper than in-house PCR for rapid identification of MTC from BACTEC MGIT 960 and BACTEC 9120 culture systems in real-time testing of AFB positive cultures.</p

Sputum microbiota profiles of treatment-naïve TB patients in Uganda before and during first-line therapy

Information on microbiota dynamics in pulmonary tuberculosis (TB) in Africa is scarce. Here, we sequenced sputa from 120 treatment-naïve TB patients in Uganda, and investigated changes in microbiota of 30 patients with treatment-response follow-up samples. Overall, HIV-status and anti-TB treatment were associated with microbial structural and abundance changes. The predominant phyla were Bacteroidetes, Firmicutes, Proteobacteria, Fusobacteria and Actinobacteria, accounting for nearly 95% of the sputum microbiota composition; the predominant genera across time were Prevotella, Streptococcus, Veillonella, Haemophilus, Neisseria, Alloprevotella, Porphyromonas, Fusobacterium, Gemella, and Rothia. Treatment-response follow-up at month 2 was characterized by a reduction in abundance of Mycobacterium and Fretibacterium, and an increase in Ruminococcus and Peptococcus; month 5 was characterized by a reduction in Tannerella and Fusobacterium, and an increase in members of the family Neisseriaceae. The microbiota core comprised of 44 genera that were stable during treatment. Hierarchical clustering of this core’s abundance distinctly separated baseline (month 0) samples from treatment follow-up samples (months 2/5). We also observed a reduction in microbial diversity with 9.1% (CI 6–14%) of the structural variation attributed to HIV-status and anti-TB treatment. Our findings show discernible microbiota signals associated with treatment with potential to inform anti-TB treatment response monitoring

Acute hypoxaemic respiratory failure in a low-income country: a prospective observational study of hospital prevalence and mortality.

INTRODUCTION: Limited data exist on the epidemiology of acute hypoxaemic respiratory failure (AHRF) in low-income countries (LICs). We sought to determine the prevalence of AHRF in critically ill adult patients admitted to a Ugandan tertiary referral hospital; determine clinical and treatment characteristics as well as assess factors associated with mortality. MATERIALS AND METHODS: We conducted a prospective observational study at the Mulago National Referral and Teaching Hospital in Uganda. Critically ill adults who were hospitalised at the emergency department and met the criteria for AHRF (acute shortness of breath for less than a week) were enrolled and followed up for 90 days. Multivariable analyses were conducted to determine the risk factors for death. RESULTS: A total of 7300 patients was screened. Of these, 327 (4.5%) presented with AHRF. The majority (60 %) was male and the median age was 38 years (IQR 27-52). The mean plethysmographic oxygen saturation (SpO2) was 77.6% (SD 12.7); mean SpO2/FiO2 ratio 194 (SD 32) and the mean Lung Injury Prediction Score (LIPS) 6.7 (SD 0.8). Pneumonia (80%) was the most common diagnosis. Only 6% of the patients received mechanical ventilatory support. In-hospital mortality was 77% with an average length of hospital stay of 9.2 days (SD 7). At 90 days after enrolment, the mortality increased to 85%. Factors associated with mortality were severity of hypoxaemia (risk ratio (RR) 1.29 (95% CI 1.15 to 1.54), p=0.01); a high LIPS (RR 1.79 (95% CI 1.79 1.14 to 2.83), p=0.01); thrombocytopenia (RR 1.23 (95% CI 1.11 to 1.38), p=0.01); anaemia (RR 1.15 (95% CI 1.01 to 1.31), p=0.03) ; HIV co-infection (RR 0.84 (95% CI 0.72 to 0.97), p=0.019) and male gender (RR 1.15 (95% CI 1.01 to 1.31) p=0.04). CONCLUSIONS: The prevalence of AHRF among emergency department patients in a tertiary hospital in an LIC was low but was associated with very high mortality. Pneumonia was the most common cause of AHRF. Mortality was associated with higher severity of hypoxaemia, high LIPS, anaemia, HIV co-infection, thrombocytopenia and being male

Microbial characteristics of dental caries in HIV positive individuals

BACKGROUND: Dental caries is a multifactorial disease that affects many people. Even though microorganisms play a crucial role in causing dental caries, diagnosis is routinely macroscopic. In order to improve early detection especially in HIV patients who are disproportionately affected, there is need to reconcile the macroscopic and microscopic characteristics of dental caries. Therefore, the aim of this study was to characterize the oral microbiota profile along the decayed, missing, filled teeth (DMFT) index using amplicon sequencing data. METHODS: Amplicon sequencing of the V6-V8 region of the 16S rRNA gene was done on DNA recovered from whole unstimulated saliva of 59 HIV positive and 29 HIV negative individuals. The microbial structure, composition and co-occurrence networks were characterized using QIIME-2, Phyloseq, Microbiome-1.9.2 and Metacoder in R. RESULTS: We characterized the oral microbiota into 2,093 operational taxonomic units (OTUs), 21 phyla and 239 genera from 2.6 million high quality sequence reads. While oral microbiota did not cluster participants into distinct groups that track with the DMFT index, we observed the following: (a) The proportion of accessory microbiota was highest in the high DMFT category while the core size (∼50% of richness) remained relatively stable across all categories. (b) The abundance of core genera such as Stomatobaculum, Peptostreptococcus and Campylobacter was high at onset of dental caries, (c) A general difference in oral microbial biomass. (d) The onset of dental caries (low DMFT) was associated with significantly lower oral microbial entropy. CONCLUSIONS: Although oral microbial shifts along the DMFT index were not distinct, we demonstrated the potential utility of microbiota dynamics to characterize oral disease. Therefore, we propose a microbial framework using the DMFT index to better understand dental caries among HIV positive people in resource limited settings

Rates of HIV-1 virological suppression and patterns of acquired drug resistance among fisherfolk on first-line antiretroviral therapy in Uganda.

OBJECTIVES: We examined virological outcomes, patterns of acquired HIV drug resistance (ADR), correlates of virological failure (VF) and acquired drug resistance among fisherfolk on first-line ART. METHODS: We enrolled 1169 adults on ART for a median duration of 6, 12, 24, 36 and ≥48 months and used a pooled VL testing approach to identify VF (VL ≥1000 copies/mL). We performed genotyping among VF cases and determined correlates of VF and ADR by logistic regression. RESULTS: The overall virological suppression rate was 91.7% and ADR was detected in 71/97 (73.2%) VF cases. The most prevalent mutations were M184V/I (53.6%) for NRTIs and K103N (39.2%) for NNRTIs. Thymidine analogue mutations were detected in 21.6% of VF cases while PI mutations were absent. A zidovudine-based ART regimen, duration on ART (≥24 months) and secondary/higher education level were significantly associated with VF. A nevirapine-based regimen [adjusted OR (aOR): 1.87; 95% CI: 0.03-0.54)] and VL ≥10000 copies/mL (aOR: 3.48; 95% CI: 1.37-8.85) were ADR correlates. The pooling strategies for VL testing with a negative predictive value (NPV) of ≥95.2% saved US $20320 (43.5%) in VL testing costs. CONCLUSIONS: We observed high virological suppression rates among these highly mobile fisherfolk; however, there was widespread ADR among those with VF at the first VL testing prior to intensive adherence counselling. Timely treatment switching and adherence support is recommended for better treatment outcomes. Adoption of pooled VL testing could be cost effective, particularly in resource-limited settings

Characteristics and outcomes of admitted patients infected with SARS-CoV-2 in Uganda

© Author(s) (or their employer(s)) 2020. Re-use permitted under CC BY-NC. No commercial re-use. See rights and permissions. Published by BMJ.Rationale: Detailed data on the characteristics and outcomes of patients with COVID-19 in sub-Saharan Africa are limited. Objective: We determined the clinical characteristics and treatment outcomes of patients diagnosed with COVID-19 in Uganda . Measurements: As of the 16 May 2020, a total of 203 cases had been confirmed. We report on the first 56 patients; 29 received hydroxychloroquine (HCQ) and 27 did not. Endpoints included admission to intensive care, mechanical ventilation or death during hospitalisation. Main results: The median age was 34.2 years; 67.9% were male; and 14.6% were 130/90 mm Hg, and 27.8% had BP of >140/90 mm Hg. Laboratory derangements were leucopenia (10.6%), lymphopenia (11.1%) and thrombocytopenia (26.3%). Abnormal chest X-ray was observed in 14.3%. No patients reached the primary endpoint. Time to clinical recovery was shorter among patients who received HCQ, but this difference did not reach statistical significance. Conclusion: Most of the patients with COVID-19 presented with mild disease and exhibited a clinical trajectory not similar to other countries. Outcomes did not differ by HCQ treatment status in line with other concluded studies on the benefit of using HCQ in the treatment of COVID-19.Publisher PDFPeer reviewe

The evolving SARS-CoV-2 epidemic in Africa: Insights from rapidly expanding genomic surveillance

INTRODUCTION Investment in Africa over the past year with regard to severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) sequencing has led to a massive increase in the number of sequences, which, to date, exceeds 100,000 sequences generated to track the pandemic on the continent. These sequences have profoundly affected how public health officials in Africa have navigated the COVID-19 pandemic. RATIONALE We demonstrate how the first 100,000 SARS-CoV-2 sequences from Africa have helped monitor the epidemic on the continent, how genomic surveillance expanded over the course of the pandemic, and how we adapted our sequencing methods to deal with an evolving virus. Finally, we also examine how viral lineages have spread across the continent in a phylogeographic framework to gain insights into the underlying temporal and spatial transmission dynamics for several variants of concern (VOCs). RESULTS Our results indicate that the number of countries in Africa that can sequence the virus within their own borders is growing and that this is coupled with a shorter turnaround time from the time of sampling to sequence submission. Ongoing evolution necessitated the continual updating of primer sets, and, as a result, eight primer sets were designed in tandem with viral evolution and used to ensure effective sequencing of the virus. The pandemic unfolded through multiple waves of infection that were each driven by distinct genetic lineages, with B.1-like ancestral strains associated with the first pandemic wave of infections in 2020. Successive waves on the continent were fueled by different VOCs, with Alpha and Beta cocirculating in distinct spatial patterns during the second wave and Delta and Omicron affecting the whole continent during the third and fourth waves, respectively. Phylogeographic reconstruction points toward distinct differences in viral importation and exportation patterns associated with the Alpha, Beta, Delta, and Omicron variants and subvariants, when considering both Africa versus the rest of the world and viral dissemination within the continent. Our epidemiological and phylogenetic inferences therefore underscore the heterogeneous nature of the pandemic on the continent and highlight key insights and challenges, for instance, recognizing the limitations of low testing proportions. We also highlight the early warning capacity that genomic surveillance in Africa has had for the rest of the world with the detection of new lineages and variants, the most recent being the characterization of various Omicron subvariants. CONCLUSION Sustained investment for diagnostics and genomic surveillance in Africa is needed as the virus continues to evolve. This is important not only to help combat SARS-CoV-2 on the continent but also because it can be used as a platform to help address the many emerging and reemerging infectious disease threats in Africa. In particular, capacity building for local sequencing within countries or within the continent should be prioritized because this is generally associated with shorter turnaround times, providing the most benefit to local public health authorities tasked with pandemic response and mitigation and allowing for the fastest reaction to localized outbreaks. These investments are crucial for pandemic preparedness and response and will serve the health of the continent well into the 21st century

High prevalence of phenotypic pyrazinamide resistance and its association with pncA gene mutations in Mycobacterium tuberculosis isolates from Uganda.

INTRODUCTION:Susceptibility testing for pyrazinamide (PZA), a cornerstone anti-TB drug is not commonly done in Uganda because it is expensive and characterized with technical difficulties thus resistance to this drug is less studied. Resistance is commonly associated with mutations in the pncA gene and its promoter region. However, these mutations vary geographically and those conferring phenotypic resistance are unknown in Uganda. This study determined the prevalence of PZA resistance and its association with pncA mutations. MATERIALS AND METHODS:Using a cross-sectional design, archived isolates collected during the Uganda national drug resistance survey between 2008-2011 were sub-cultured. PZA resistance was tested by BACTEC Mycobacterial Growth Indicator Tube (MGIT) 960 system. Sequence reads were downloaded from the NCBI Library and bioinformatics pipelines were used to screen for PZA resistance-conferring mutations. RESULTS:The prevalence of phenotypic PZA resistance was found to be 21%. The sensitivity and specificity of pncA sequencing were 24% (95% CI, 9.36-45.13%) and 100% (73.54% - 100.0%) respectively. We identified four mutations associated with PZA phenotypic resistance in Uganda; K96R, T142R, R154G and V180F. CONCLUSION:There is a high prevalence of phenotypic PZA resistance among TB patients in Uganda. The low sensitivity of pncA gene sequencing confirms the already documented discordances suggesting other mechanisms of PZA resistance in Mycobacterium tuberculosis