Analytical performances of the Xpert MTB/RIF assay using stool specimens to improve the diagnosis of pulmonary tuberculosis in Burkina Faso, a tuberculosis endemic country.

Timely diagnosis of Pulmonary Tuberculosis (PTB) is associated with good prognosis, but remains difficult in primary healthcare facilities and particularly in children and patients living with HIV. The aim of this study was to compare the GeneXpert ® MTB/RIF assay (Xpert) performed using a stool sample (3-5 g) and using the first Respiratory Tract Sample (RTS; i.e., sputum, bronchoalveolar or gastric aspirate; as normally done) concomitantly collected from 119 patients with suspected PTB to improve PTB diagnosis in Burkina Faso, a high tuberculosis burden country with limited resources. Overall, microbiological, microscopic and molecular analysis of the 119 first RTS and 119 stool specimens led to Mycobacterium tuberculosis complex detection in 28 patients (23 positive RTS cultures and 5 negative RTS cultures-RTS Xpert positive). When using the 28 clinical confirmed cases as reference standard, the sensitivities of the stool-based and RTS-based Xpert assays were not different (24/28, 85.7%, versus 26/28, 92.86%; p > 0.30), and 22 results were fully concordant. Considering the first RTS culture as the gold standard, the sensitivities of the stool-based and RTS-based Xpert assays to detect PTB in patients with positive RTS culture were 100% (23/23) and 91.3% (21/23), respectively (p >0.05). The stool-based Xpert assay specificity for excluding PTB was 99% (95/96) (compared with 95%, 91/96, when using RTS) and its negative and positive predictive values were 100% (95/95) and 96% (23/24), respectively. Compared with the 23 positive RTS cultures, the incremental yield rates of the RTS-based and stool-based Xpert assays were 4.2% (5/119) and 0.84% (1/119), respectively. Overall, our findings support using the stool-based Xpert assay as an alternative method for earlier PTB diagnosis, when RTS are difficult to obtain.

Seroprevalence of SARS-CoV-2 IgG and associated factors among people living with HIV over the first 12 months following the outbreak of COVID-19 in Burkina Faso, a sub-Saharan African country.

OBJECTIVE: This study aimed to evaluate the seroprevalence of anti-SARS-CoV-2 IgG and factors associated with the infection among PLWHIV over the first 12 months following the outbreak of COVID-19 in Burkina Faso. DESIGN: A retrospective cross-sectional study of plasma samples collected from March 9, 2020, and March 8, 2021, at the outpatient HIV referral center, before the introduction of the SARS-CoV-2 vaccine in Burkina Faso. METHODS: Anti-SARS-CoV-2 IgG were detected in plasma using DS-ЕIA-ANTI-SARS-CoV-2-G (S) kit. Logistic regressions were used to compare SARS-CoV-2 specific immune responses between groups and within subgroups. RESULTS AND DISCUSSION: A total of 419 plasma were subjected to serological diagnosis. None of the participants was vaccinated against COVID-19 during the period of sample collection, and 130 samples were positive for anti-SARS-CoV-2 IgG, giving a prevalence of 31.0% (95% CI 26.6-35.7). The median CD4 cell count was 661 cells/µL (IQR,422-928). Retailers had half the risk of being infected compared to housemaids with an OR of 0.49 (p = 0.028, 95% CI 0.26-0.91). Likewise, the risk of infection was 1.69 times higher in patients on integrase inhibitors compared to that of patients on non-nucleoside reverse transcriptase inhibitors (p = 0.020, 95% CI 1.09-2.63). CONCLUSION: Our study reveals a high seroprevalence among PLWHIV to SARS-CoV-2 during the first year of the pandemic. In addition, PLWHIV on integrase inhibitors are 1.69 times more likely to be infected than PLWHIV on non-nucleoside inhibitors, and this observation remains an intriguing topic that still needs to be clarified.

Evaluation of the impact of COVID-19 in people coinfected with HIV and/or tuberculosis in low-income countries: study protocol for mixed methods research in Burkina Faso.

BACKGROUND: An issue of particular concern is the impact of the 2019 novel coronavirus (2019 nCOV) on the people coinfected with the Human Immuno-deficiency Virus (HIV) and/or tuberculosis (TB). Unfortunately, this interaction has not been well explored in African despite the large proportion of these risk populations living with HIV and/or patients and/or tuberculosis (TB) in the African region. This study aims to design a research protocol for assessment of the impact of coronavirus disease 2019 (COVID-19) on these risk populations in response to COVID-19 strategic plans in Burkina Faso by generating serological, epidemiological, virological, clinical and socio-anthropological evidence-based data. METHODS: A multidisciplinary research will be conducted in the city of Bobo-Dioulasso, Burkina Faso using mixed methods. Data will be collected from a cohort of people living with HIV and/or TB patients in the city (i) to determine the proportion of people with specific antibodies against the severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) using retrospective data ; (ii) to determine the proportion of people infected with Covid-19 and the dynamics of viral loads and antibodies in these people based on prospective data; (iii) to identify circulating SARS-COV-2 variants and novel biomarkers using prospective data ; (iv) to analyze perceptions, community experiences and response strategies during the public health emergencies imposed by COVID-19 through a qualitative study. DISCUSSION: This study will generate factual and comprehensive data that will contribute in improving response strategies to COVID-19 and the other possible emerging diseases with keen interest on the risk populations living with HIV and/or TB infected patients.

Planctomycetes as Host-Associated Bacteria: A Perspective That Holds Promise for Their Future Isolations, by Mimicking Their Native Environmental Niches in Clinical Microbiology Laboratories.

Traditionally recognized as environmental bacteria, Planctomycetes have just been linked recently to human pathology as opportunistic pathogens, arousing a great interest for clinical microbiologists. However, the lack of appropriate culture media limits our future investigations as no Planctomycetes have ever been isolated from patients' specimens despite several attempts. Several Planctomycetes have no cultivable members and are only recognized by 16S rRNA gene sequence detection and analysis. The cultured representatives are slow-growing fastidious bacteria and mostly difficult to culture on synthetic media. Accordingly, the provision of environmental and nutritional conditions like those existing in the natural habitat where yet uncultured/refractory bacteria can be detected might be an option for their potential isolation. Hence, we systematically reviewed the various natural habitats of Planctomycetes, to review their nutritional requirements, the physicochemical characteristics of their natural ecological niches, current methods of cultivation of the Planctomycetes and gaps, from a perspective of collecting data in order to optimize conditions and the protocols of cultivation of these fastidious bacteria. Planctomycetes are widespread in freshwater, seawater, and terrestrial environments, essentially associated to particles or organisms like macroalgae, marine sponges, and lichens, depending on the species and metabolizable polysaccharides by their sulfatases. Most Planctomycetes grow in nutrient-poor oligotrophic environments with pH ranging from 3.4 to 11, but a few strains can also grow in quite nutrient rich media like M600/M14. Also, a seasonality variation of abundance is observed, and bloom occurs in summer-early autumn, correlating with the strong growth of algae in the marine environments. Most Planctomycetes are mesophilic, but with a few Planctomycetes being thermophilic (50°C to 60°C). Commonly added nutrients are N-acetyl-glucosamine, yeast-extracts, peptone, and some oligo and macro-elements. A biphasic host-associated extract (macroalgae, sponge extract) conjugated with a diluted basal medium should provide favorable results for the success of isolation in pure culture.

Escherichia coli Culture Filtrate Enhances the Growth of Gemmata spp.

BACKGROUND: Planctomycetes bacteria are known to be difficult to isolate, we hypothesized this may be due to missing iron compounds known to be important for other bacteria. We tested the growth-enhancement effect of complementing two standard media with Escherichia coli culture filtrate on two cultured strains of Gemmata spp. Also, the acquisition of iron by Gemmata spp. was evaluated by measuring various molecules involved in iron metabolism. MATERIALS AND METHODS: Gemmata obscuriglobus and Gemmata massiliana were cultured in Caulobacter and Staley's medium supplemented or not with E. coli culture filtrate, likely containing siderophores and extracellular ferrireductases. We performed iron metabolism studies with FeSO4, FeCl3 and deferoxamine in the cultures with the E. coli filtrate and the controls. RESULTS AND DISCUSSION: The numbers of G. obscuriglobus and G. massiliana colonies on Caulobacter medium or Staley's medium supplemented with E. coli culture filtrate were significantly higher than those on the standard medium (p < 0.0001). Agar plate assays revealed that the Gemmata colonies near E. coli colonies were larger than the more distant colonies, suggesting the diffusion of unknown growth promoting molecules. The inclusion of 10-4 to 10-3 M FeSO4 resulted in rapid Gemmata spp. growth (4-5 days compared with 8-9 days for the controls), suggesting that both species can utilize FeSO4 to boost their growth. In contrast, deferoxamine slowed down and prevented Gemmata spp. growth. Further studies revealed that the complementation of Caulobacter medium with E. coli culture filtrate and 10-4 M FeSO4 exerted a significant growth-enhancement effect compared with that obtained with Caulobacter medium supplemented with E. coli culture filtrate alone (p < 0.0122). Moreover, the intracellular iron concentrations in G. obscuriglobus and G. massiliana cultures in iron-depleted broth supplemented with the E. coli filtrate were 0.63 ± 0.16 and 0.78 ± 0.12 µmol/L, respectively, whereas concentrations of 1.72 ± 0.13 and 1.56± 0.11 µmol/L were found in the G. obscuriglobus and G. massiliana cultures grown in broth supplemented with the E. coli filtrate and FeSO4. The data reported here indicated that both E. coli culture filtrate and FeSO4 act as growth factors for Gemmata spp. via a potentiation mechanism.

Improved culture of fastidious Gemmata spp. bacteria using marine sponge skeletons.

Gemmata are Planctomycetes bacteria recalcitrant to traditional cultivation in the clinical microbiology laboratory and they have been seldom documented in patients. Based on previously known relationships of Planctomycetes with marine sponges, we designed a new culture medium A incorporating marine sponge skeleton of Spongia sp. to the standard culture medium; and culture medium B incorporating Spongia sp. skeleton heat aqueous filtrate into medium A; and inoculating the three culture media (standard, A and B) with Gemmata obscuriglobus DSM 5831T and Gemmata massiliana DSM 26013T in the presence of negative controls. Cultures were observed by naked eyes for 7 days and bacterial growth was quantified by microscopic observations and culture-based enumerations. Macroscopic observations at day-3 revealed a pink bacterial pellet in medium B tubes while standard medium tubes remained limpid until day-8. Growing Gemmata spp. bacteria in medium A yielded air bubbles released by bacterial respiration, whereas control tubes remained bubble-free. The number of colonies in standard medium (1.363 ± 115 for G. obscuriglobus, 1.288 ± 83 for G. massiliana) was significantly lower than those counted from medium B (2.552 ± 128 for G. obscuriglobus, 1.870 ± 112 for G. massiliana) and from medium A (2.851 ± 137 for G. obscuriglobus, 2.035 ± 163 for G. massiliana) (p < 0.10-4) at day-2 incubation. At day-3 incubation, the number of colonies counted from supplemented media A and B increased up to one log than those counted from the control medium (p < 0.10-4). Along the following day-4-7 incubation, the number of colonies counted from media A and B remained significantly higher compared to standard medium (p < 0.10-4). These data indicate that incorporation of spongin-based marine sponge skeleton and heat aqueous filtrate of sponge skeleton significantly improved growth of Gemmata spp. bacteria. These observations pave the way towards improved isolation and culture of Gemmata spp. from environmental and clinical specimens.

Co-culture models illustrate the digestion of Gemmata spp. by phagocytes.

Gemmata spp. bacteria thrive in the same aquatic environments as free-living amoebae. DNA-based detection of Gemmata spp. sequences in the microbiota of the human digestive tract and blood further questioned the susceptibility of Gemmata spp. to phagocytes. Here, Gemmata obscuriglobus and Gemmata massiliana were co-cultured with the amoebae Acanthamoeba polyphaga, Acanthamoeba castellanii, Acanthamoeba griffini and THP-1 macrophage-like phagocytes. All experiments were performed in five independant replicates. The ratio amoeba/bacteria was 1:20 and the ratio THP-1/bacteria was 1:10. After a 2-hour co-culture, extracellular bacteria were killed by kanamycin or amikacin and eliminated. The intracellular location of Gemmata bacteria was specified by confocal microscopy. Microscopic enumerations and culture-based enumerations of colony-forming units were performed at T = 0, 1, 2, 3, 4, 8, 16, 24, 48 and 72 hours post-infection. Then, Gemmata bacteria were engulfed into the phagocytes' cytoplasmic vacuoles, more than (98 ± 2)% of Gemmata bacteria, compared to controls, were destroyed by phagocytic cells after a 48-h co-culture according to microscopy and culture results, and no positive culture was observed at T = 72-hours. Under our co-culture conditions, Gemmata bacteria were therefore susceptible to the environmental and host phagocytes here investigated. These data suggest that these Acanthamoeba species and THP-1 cells cannot be used to isolate G. massiliana and G. obscuriglobus under the co-culture conditions applied in this study. Although the THP-1 response can point towards potential responses that might occur in vivo, these responses should first bevalidated by in vivo studies to draw definite conclusions.