Combination of serological and cytokine release assays for improved diagnosis of childhood tuberculosis in Zambia (PROMISE-TB).

OBJECTIVES: The diagnostic gaps for childhood tuberculosis (TB) remain considerable in settings with high TB incidence and resource constraints. We established and evaluated the performance of a scoring system based on a combination of serological tests and T-cell cytokine release assays, chosen for their ability to detect immune responses indicative of TB, in a context of high prevalence of pediatric HIV infection. METHODS: We enrolled 628 consecutive children aged ≤15 years, admitted for TB suspicion. Multiple cytokine levels in QuantiFERON Gold In-Tube supernatants and antigen 85B (Ag85B) antibodies were assessed in children who tested positive with either Xpert TB or mycobacterial culture. The results were compared with those of control children. RESULTS: Among the biomarkers most strongly associated with TB, random forest classification analysis selected Ag85B antibodies, interleukin-2/interferon-γ ratio, and monokine induced by interferon-γ for the scoring system. The receiver operating characteristic curve derived from our scoring system showed an area under the curve of 0.95 (0.91-0.99), yielding 91% sensitivity and 88% specificity. The internal bootstrap validation gave the following 95% confidence intervals for the score performance: sensitivity 71%-97% and specificity 79%-99%. CONCLUSIONS: This study suggests that supplementing the QuantiFERON assay with a combination of serological and T-cell markers could enhance childhood TB screening regardless of HIV status and age. Further validation among the target population is necessary to confirm the performance of this scoring system.

Development and field evaluation in African and Asian countries of an hepatitis B virus PCR on open polyvalent platforms to determine treatment eligibility: results from the "Agence Nationale de Recherche sur le Sida et les hépatites" 12327 study.

OBJECTIVES: Widespread testing and treatment are essential to eliminate hepatitis B virus (HBV) infection as a public health concern. However, in resource-limited countries, access to HBV PCR is limited. In this study, we developed a quantitative HBV PCR assay on open molecular platforms and evaluate its performance in diagnosing clinically significant HBV DNA thresholds as defined by the WHO (2000 IU/mL, 20 000 IU/mL, and 200 000 IU/mL). METHODS: We implemented our HBV PCR test in seven African and Asian countries and France, using either an in-house laboratory method or a European conformity for in vitro diagnostic (CE-IVD) marked version of the PCR (Generic HBV Charge Virale, Biocentric). Results were compared with reference tests (Roche Cobas AmpliPrep/Cobas TaqMan and Abbott RealTime on Abbott m2000). RESULTS: There was a good agreement between the HBV DNA results of 1015 samples tested by the PCR on open polyvalent platforms and the results from reference tests (mean difference (bias ± standard deviation [SD]): -0.3 ± 0.7 log10 IU/mL and -0.2 ± 0.9 log10 IU/mL when compared with Roche and Abbott tests, respectively). Kappa-Cohen agreements between the HBV PCR on open polyvalent platforms and the Roche/Abbott assays appeared almost perfect for HBV DNA levels ranged from >20 000 to 200 000 IU/mL and >200 000 IU/mL, substantial and moderate for HBV DNA levels ranged from 2000 to 20 000 IU/mL when compared with Abbott and Roche, respectively. The assay's performance was consistent across genotypes A, B, C, D, and E. DISCUSSION: This field evaluation showed that our HBV PCR test is a valuable alternative to proprietary PCR systems. PCR assays on open platforms contribute to expanding clinical laboratory solutions for diagnosing individuals who meet the viral load criteria for antiviral therapy (>20 000 IU/mL) and mother-to-child prophylaxis (>200 000 IU/mL).

Evolution and spread of Plasmodium falciparum mutations associated with resistance to sulfadoxine-pyrimethamine in central Africa: a cross-sectional study.

BACKGROUND: Efficacy of sulfadoxine-pyrimethamine, the malaria chemoprophylaxis used in pregnant women, and in children when combined with amodiaquine, is threatened by the accumulation of mutations in the Plasmodium falciparum dihydropteroate synthase (pfdhps) and dihydrofolate reductase (pfdhfr) genes. Data on the prevalence of resistant alleles in central Africa and the new pfdhps I431V mutation, particularly associated with other mutations to form the pfdhps vagKgs allele, are scarce. We explored the frequency and geographical distribution of pfdhps and pfdhfr mutations in central Africa in 2014-18, and assessed the evolutionary origin of the vagKgs allele. METHODS: Samples were collected at 18 health-care centres in seven countries (Angola, Cameroon, Central African Republic, Democratic Republic of the Congo, Gabon, Nigeria, and Republic of the Congo) from patients who showed possible symptoms of malaria between March 1, 2014, and Oct 31, 2018. Samples that were positive for P falciparum were transported to a laboratory in Toulouse, France, and genotyped. The frequency of pfdhfr and pfdhps mutations was studied in 1749 samples. Microsatellites in pfdhps flanking regions and whole-genome analysis compared with parasite genomes from the data-sharing network MalariaGEN were performed on samples carrying the vagKgs allele. FINDINGS: Mapping of the prevalence of single nucleotide polymorphisms and corresponding alleles of pfdhfr and pfdhps showed a substantial spread of alleles associated with sulfadoxine-pyrimethamine resistance in central Africa during the 2014-18 period, especially an increase going west to east in pfdhps alleles carrying the K540E and A581G mutations. A high prevalence of the pfdhps I431V mutation was observed in Cameroon (exceeding 50% in the northern region) and Nigeria. Genomic analysis showed a recent African emergence and a clonal expansion of the most frequent pfdhps vagKgs allele. INTERPRETATION: Reduced sulfadoxine-pyrimethamine efficacy due to increased resistance is a worrying situation, especially because the malaria transmission level is high in central Africa. Although the resistance phenotype remains to be confirmed, the emergence and spread of the vagKgs allele in west and central Africa could challenge the use of sulfadoxine-pyrimethamine. FUNDING: Toulouse Institute for Infectious and Inflammatory Diseases.

Burkholderia cepacia meningitis in the Central African Republic.

Burkholderia cepacia causes frequent infections in immunocompromised and hospitalized patients, with a significant mortality rate. This bacterial species has also been associated with epidemic outbreaks due to contamination of antiseptic solutions and parenteral and nebulized medications. In 2016, in the town of Bongonon in the north of the Central African Republic (CAR), a three-year-old boy with febrile meningeal syndrome (fever, neck stiffness and altered general condition) was admitted for a medical consultation provided by the nongovernmental organization MSF-Spain. On 20 March 2016, a sample of the boy's cerebrospinal fluid was sent to the Bacteriology Laboratory of the Pasteur Institute of Bangui for analysis. Conventional bacteriology showed that the isolate was a Gram-negative bacillus, which was identified as B. cepacia by using API 20 NE, with 99.9%confidence. In addition, the strain presented an acquired resistance to ticarcillin-clavulanate, ceftazidime and imipenem but remained susceptible to cotrimoxazole. As B. cepacia had never previously been isolated from cerebrospinal fluid in Africa, we chose to identify the strain by 16S rRNA gene sequencing. The molecular data showed that the isolate belonged to B. cepacia group. This is the first report of a case of meningitis caused by B. cepacia in CAR and developing countries.

Emergence of Neisseria meningitidis Serogroup W, Central African Republic, 2015-2016.

We analyzed data from the 2015 and 2016 meningitis epidemic seasons in Central African Republic as part of the national disease surveillance. Of 80 tested specimens, 66 belonged to meningococcal serogroup W. Further analysis found that 97.7% of 44 isolates belonged to the hyperinvasive clonal complex sequence type 11.

Evidence of human leptospirosis cases in a cohort of febrile patients in Bangui, Central African Republic: a retrospective study, 2012-2015.

BACKGROUND: In spite of a local favorable environment, leptospirosis has never been described in Central African Republic so far mainly because of the weakness of diagnostic tests and differential diagnostic strategy for febrile jaundice cases negative for yellow fever virus. Here we bring a complementary insight to conclusions of Gadia CLB et al. regarding the presence of leptospirosis in Central African Republic in YFV-negative febrile icteric patients. METHODS: Our study included 497 individuals presenting with fever and jaundice but negative for yellow fever infection, retrospectively selected from the national surveillance biobank for yellow fever in Institut Pasteur de Bangui, Central African Republic. A combination of serological (ELISA, agglutination) and molecular biology techniques (quantitative real-time polymerase chain reaction) was used to identify Leptospira or the patient's immune response to the bacteria. Statistical analyses were done using the non parametric Mann-Withney U test with a 5% statistical threshold. RESULTS: ELISA test results showed 46 positive serum samples while 445 were negative and 6 remains equivocal. In addition, the reference microscopic agglutination test for leptospirosis diagnostic confirmed that 7 out of 32 samples tested were positive. Unfortunately, all 497 serum samples tested for leptospirosis were negative using the molecular techniques. CONCLUSIONS: Unlike Gadia et al., we confirmed that leptospirosis is circulating in Central African Republic and therefore may be responsible for some of the unexplained cases of febrile jaundice in the country. Thus, leptospirosis needs to be investigated to improve identification of aetiological pathogens. Our study also suggests a need to improve sample transportation and storage conditions.

Stunted childhood growth is associated with decompartmentalization of the gastrointestinal tract and overgrowth of oropharyngeal taxa.

Linear growth delay (stunting) affects roughly 155 million children under the age of 5 years worldwide. Treatment has been limited by a lack of understanding of the underlying pathophysiological mechanisms. Stunting is most likely associated with changes in the microbial community of the small intestine, a compartment vital for digestion and nutrient absorption. Efforts to better understand the pathophysiology have been hampered by difficulty of access to small intestinal fluids. Here, we describe the microbial community found in the upper gastrointestinal tract of stunted children aged 2-5 y living in sub-Saharan Africa. We studied 46 duodenal and 57 gastric samples from stunted children, as well as 404 fecal samples from stunted and nonstunted children living in Bangui, Central African Republic, and in Antananarivo, Madagascar, using 16S Illumina Amplicon sequencing and semiquantitative culture methods. The vast majority of the stunted children showed small intestinal bacterial overgrowth dominated by bacteria that normally reside in the oropharyngeal cavity. There was an overrepresentation of oral bacteria in fecal samples of stunted children, opening the way for developing noninvasive diagnostic markers. In addition, Escherichia coli/Shigella sp. and Campylobacter sp. were found to be more prevalent in stunted children, while Clostridia, well-known butyrate producers, were reduced. Our data suggest that stunting is associated with a microbiome "decompartmentalization" of the gastrointestinal tract characterized by an increased presence of oropharyngeal bacteria from the stomach to the colon, hence challenging the current view of stunting arising solely as a consequence of small intestine overstimulation through recurrent infections by enteric pathogens.

Identifying the etiology and pathophysiology underlying stunting and environmental enteropathy: study protocol of the AFRIBIOTA project.

BACKGROUND: Globally one out of four children under 5 years is affected by linear growth delay (stunting). This syndrome has severe long-term sequelae including increased risk of illness and mortality and delayed psychomotor development. Stunting is a syndrome that is linked to poor nutrition and repeated infections. To date, the treatment of stunted children is challenging as the underlying etiology and pathophysiological mechanisms remain elusive. We hypothesize that pediatric environmental enteropathy (PEE), a chronic inflammation of the small intestine, plays a major role in the pathophysiology of stunting, failure of nutritional interventions and diminished response to oral vaccines, potentially via changes in the composition of the pro- and eukaryotic intestinal communities. The main objective of AFRIBIOTA is to describe the intestinal dysbiosis observed in the context of stunting and to link it to PEE. Secondary objectives include the identification of the broader socio-economic environment and biological and environmental risk factors for stunting and PEE as well as the testing of a set of easy-to-use candidate biomarkers for PEE. We also assess host outcomes including mucosal and systemic immunity and psychomotor development. This article describes the rationale and study protocol of the AFRIBIOTA project. METHODS: AFRIBIOTA is a case-control study for stunting recruiting children in Bangui, Central African Republic and in Antananarivo, Madagascar. In each country, 460 children aged 2-5 years with no overt signs of gastrointestinal disease are recruited (260 with no growth delay, 100 moderately stunted and 100 severely stunted). We compare the intestinal microbiota composition (gastric and small intestinal aspirates; feces), the mucosal and systemic immune status and the psychomotor development of children with stunting and/or PEE compared to non-stunted controls. We also perform anthropological and epidemiological investigations of the children's broader living conditions and assess risk factors using a standardized questionnaire. DISCUSSION: To date, the pathophysiology and risk factors of stunting and PEE have been insufficiently investigated. AFRIBIOTA will add new insights into the pathophysiology underlying stunting and PEE and in doing so will enable implementation of new biomarkers and design of evidence-based treatment strategies for these two syndromes.

B cells response directed against Cut4 and CFP21 lipolytic enzymes in active and latent tuberculosis infections.

BACKGROUND: Better understanding of the immune response directed against Mycobacterium tuberculosis (Mtb) is critical for development of vaccine strategies and diagnosis tests. Previous studies suggested that Mtb enzymes involved in lipid metabolism, are associated with persistence and/or reactivation of dormant bacilli. METHODS: Circulating antibodies secreting cells (ASCs), memory B cells, and antibodies directed against Cut4 (Rv3452) and CFP21 (Rv1984c) antigens were explored in subjects with either active- or latent-tuberculosis (LTB), and in Mtb-uninfected individuals. RESULTS: Circulating anti-Cut4 ASCs were detected in 11/14 (78.6%) subjects from the active TB group vs. 4/17 (23.5%) from the LTB group (p = 0.001). Anti-CFP21 ASCs were found in 11/14 (78.6%) active TB vs. in 5/17 (29.4%) LTB cases (p = 0.01). Circulating anti-Cut4 and anti-CFP21 ASCs were not detected in 38 Mtb uninfected controls. Memory B cells directed against either Cut4 or CFP21 were identified in 8/11 (72.7%) and in 9/11 (81.8%) subjects with LTB infection, respectively, and in 2/6 Mtb uninfected individuals (33.3%). High level of anti-Cut4 and anti-CFP21 IgG were observed in active TB cases. CONCLUSION: Circulating IgG SCs directed against Cut4 or CFP21 were mostly detected in patients presenting an active form of the disease, suggesting that TB reactivation triggers an immune response against these two antigens.

Semi-quantitative real-time PCR: A useful approach to identify persons with low replicative chronic hepatitis B.

Antiviral therapy can be avoided during the low replicative phase of chronic Hepatitis B virus (HBV) infection which is characterized notably by HBV DNA concentration below 2000IU/ml. Simplified diagnostic tests can improve access to HBV DNA monitoring in resource-limited settings. The capacity of a new semi-quantitative real-time PCR approach based on sample-to-standard relative detection of the target to discriminate samples with HBV DNA levels above or below the clinical threshold of 2000IU/ml was compared to a quantitative assay (Roche CobasAmpliPrep/CobasTaqMan HBV Test v2.0). The semi-quantitative assay correctly identified 40/40 (100%) low replicative HBV DNA patients and 58/61 (95%) samples from HBV-infected subjects with moderate/high levels of viral DNA. Our results suggested that this alternative PCR test is efficient to guide therapeutic decision based on identification of low replicative HBV infection from all of the chronic hepatitis B carriers requiring treatment, and may be useful in resource-limited settings where the vast majority of cases live.

Subclinical mastitis occurs frequently in association with dramatic changes in inflammatory/anti-inflammatory breast milk components.

BACKGROUND: Subclinical mastitis (SCM) is a frequent, but poorly characterized entity that may influence immune development of breastfed infants. Mechanisms driving the emergence of SCM and changes in immunological content of human milk during SCM remain to be explored. In this study, the breast milk environment was to describe during SCM. METHODS: One hundred and ten samples of mature breast milk were collected from 44 healthy, HIV-negative mothers, included in a large infant feeding intervention cohort (ANRS 1271/Vertical Transmission Study). Immune markers related to inflammatory/anti-inflammatory balances and secreted in response to bacterial exposure were explored in SCM breast milk samples (Na/K ratio > 1) and compared to non-SCM controls. RESULTS: SCM was observed in 23% of women (95% confidence interval (CI): 21-24) and associated with higher levels of inflammatory markers (ß2 microgobulin, PS100A9, TNF-α, IL-6, IL-8, IL-17, and RANTES) and Th1-related cytokines (IL-2R, IL-12p40/70, IFN-α, IFN-γ, CXCL-9, andIP-10). High levels of factors secreted in response to bacteria and lipopolysaccharide (LPS) exposure were observed in SCM breast milk samples (MIP-1α, MIP-1ß, LPS binding protein, α-defensins, and antileukoproteinase 1). CONCLUSION: SCM is associated with important changes in breast milk microenvironment, with a proinflammatory/Th1-cytokine predominant profile. During SCM, cytokine imbalances in breast milk may have a notable influence on mucosal immune system of the infant early in life.

High IFN-γ Release and Impaired Capacity of Multi-Cytokine Secretion in IGRA Supernatants Are Associated with Active Tuberculosis.

Interferon gamma (IFN-γ) release assays (IGRAs) detect Mycobacterium tuberculosis (Mtb) infection regardless of the active (ATB) or latent (LTBI) forms of tuberculosis (TB). In this study, Mtb-specific T cell response against region of deletion 1 (RD1) antigens were explored by a microbead multiplex assay performed in T-SPOT TB assay (T-SPOT) supernatants from 35 patients with ATB and 115 patients with LTBI. T-SPOT is positive when over 7 IFN-γ secreting cells (SC)/250 000 peripheral blood mononuclear cells (PBMC) are enumerated. However, over 100 IFN-γ SC /250 000 PBMC were more frequently observed in the ATB group compared to the LTBI group. By contrast, lower cytokine concentrations and lower cytokine productions relative to IFN-γ secretion were observed for IL 4, IL-12, TNF-α, GM-CSF, Eotaxin and IFN-α when compared to LTBI. Thus, high IFN-γ release and low cytokine secretions in relation with IFN-γ production appeared as signatures of ATB, corroborating that multicytokine Mtb-specific response against RD1 antigens reflects host capacity to contain TB reactivation. In this way, testing cytokine profile in IGRA supernatants would be helpful to improve ATB screening strategy including immunologic tests.

Cytokine Response Associated with Hepatitis C Virus Clearance in HIV Coinfected Patients Initiating Peg Interferon-α Based Therapy.

BACKGROUND: Treatment of hepatitis C virus (HCV) infection based on peginterferon-α (pegIFNα) and ribavirin induces important changes in cytokine release and T cell activation. OBJECTIVE: Immune response to pegIFNα-ribavirin therapy was explored in patients coinfected by HCV and HIV. METHODS: Concentrations of 25 cytokines and CD8(+) T cell activation were monitored in HCV/HIV coinfected patients classified as sustained virological responders (SVR, n=19) and non-responders (NR, n=11). RESULTS: High pretreatment concentrations of IP-10 (CXCL-10) and MCP-1 (CCL-2) were associated with a poor anti-HCV response. PegIFNα-ribavirin therapy increased CD8(+) T cell activation and induced significant changes in levels of eleven cytokines related to both Th1 and Th2 responses in SVR (IL-1ß, IL-1RA, IL-4, IL-5, IL-6, IL-7, IL-12p40/70, IL-13, IP-10, eotaxin, MCP-1) but of only six cytokines in NR (IL-1ß, IL-2, IL-5, IL-12p40/70, IL-13, eotaxin). The highest rise in MIP-1ß and MCP-1 levels was observed four weeks after anti-HCV treatment initiation in SVR compared to NR (p=0.002 and p=0.03, respectively), whereas a decrease in IL-8 concentration was associated with treatment failure (p= 0.052). CONCLUSIONS: Higher and broader cytokine responses to pegIFNα-ribavirin therapy were observed in SVR patients compared to NR. Changes in IL-8, MIP-1ß, and MCP-1 serum concentrations may be associated with efficacy of pegIFNα- and ribavirin-based therapies in patients coinfected by HCV and HIV.

Close relationship between immunoglobulin secreting-cells and Epstein-Barr virus reservoir in patients infected with HIV.

Individuals infected with HIV have higher circulating Epstein-Barr virus (EBV) DNA load compared to healthy carriers. This study investigated whether level of spontaneous immunoglobulin secreting cells, one of the major hallmarks of HIV infection, is associated with an increase of EBV DNA load in PBMCs and the spontaneous EBV lytic cycle ex vivo in patients infected with HIV. Spontaneous virus production by cells infected with EBV and EBV DNA loads in PBMCs from which CD8(+) T-cells were removed were measured in 20 HIV-aviremic and 14 HIV-viremic patients. The number of circulating immunoglobulin-secreting cells (Ig-SCs) and CD8(+) T-lymphocyte activation were also investigated. Patients with detectable HIV RNA in plasma exhibited higher spontaneous ex vivo EBV secretion and higher levels of EBV DNA in PBMCs than their aviremic counterparts. In the two groups observed, a positive correlation was found between PBMCs EBV DNA viral load and Ig-SCs, CD38(bright) expression on CD8(+) T-cells and EBV DNA load in cell culture supernatants. These findings suggest that B-cell polyclonal activation and B-cell terminal differentiation into Ig-SCs may fuel EBV DNA reservoir and promote EBV production ex vivo in patients infected with HIV.

In vitro infection of primary human hepatocytes by HCV-positive sera: insights on a highly relevant model.

OBJECTIVE: Adult primary human hepatocytes (PHHs) support the complete infection cycle of natural HCV from patients' sera. The molecular details underlying sera infectivity towards these cells remain largely unknown. Therefore, we sought to gain a deeper comprehension of these features in the most physiologically relevant culture system. DESIGN: Using kinetic experiments, we defined the optimal conditions to infect PHH and explored the link between cell organisation and permissivity. Based on their infectivity, about 120 sera were classified in three groups. Concentration of 52 analytes was measured in 79 selected sera using multiplexed immunobead-based analyte profiling. RESULTS: PHH permissivity towards HCV infection negatively correlated with cell polarisation and formation of functional bile canaliculi. PHH supported HCV replication for at least 2 weeks with de novo virus production. Depending on their reactivity, sera could be classified in three groups of high, intermediate or low infectivity toward PHH. Infectivity could not be predicted based on the donors' clinical characteristics, viral load or genotype. Interestingly, highly infectious sera displayed a specific cytokine profile with low levels of most of the 52 tested analytes. Among them, 24 cytokines/growth factors could impact hepatocyte biology and infection efficiency. CONCLUSIONS: We identified critical factors leading to efficient PHH infection by HCV sera in vitro. Overall, we showed that this cellular model provides a useful tool for studying the mechanism of HCV infection in its natural host cell, selecting highly infectious isolates, and determining the potency of drugs towards various HCV strains.

Increased T-cell activation and Th1 cytokine concentrations prior to the diagnosis of B-cell lymphoma in HIV infected patients.

BACKGROUND: Despite the use of combined antiretroviral therapy, HIV-infected individuals have a higher risk of developing B-cell lymphoma compared to the general population. We aim to explore whether lymphocyte activation, increase in Th1 response as well as markers of EBV reactivation, may precede lymphoma diagnosis. METHODS: Thirteen cases and 26 controls matched on CD4(+) T-cell count and HIV plasma viral load were identified. Samples were collected 0 to 5 years prior to B-cell lymphoma diagnosis. Seven out of 13 (54 %) and 16/26 (61.5 %) of cases and controls were receiving antiretroviral therapy at the time of sampling, respectively. CD8(+) T-cell activation and Th1 cytokine concentrations were measured before lymphoma onset, together with IgG antibodies directed against viral capsid antigen (VCA) and serum levels of EBV DNA. RESULTS: A higher level of CD8(+) T-cell activation was observed in patients developing lymphoma. Four out of seven Th1 cytokine serum concentrations were significantly higher in patients with lymphoma than in the control group: IL-2R, IL-12p40/70, IFN-γ-inducible protein 10 (IP-10) and monokine induced by IFN-γ (MIG). Anti-VCA IgG level were significantly higher in cases than in controls. Four cases (30 %) but no controls had detectable EBV DNA in serum. CONCLUSION: A higher level of T-cell activation, Th1 cytokine serum concentration and markers of EBV replication, preceded B-cell lymphoma diagnosis. This may suggest that viral antigen stimulation is associated with the genesis of lymphoma in HIV-infected patients.

Thalidomide for steroid-dependent immune reconstitution inflammatory syndromes during AIDS.

Management of relapsing or refractory immune reconstitution inflammatory syndromes (IRISs) despite corticosteroid therapy is yet to be defined. We describe three HIV-infected patients with corticosteroid-dependent and life-threatening paradoxical immune reconstitution inflammatory syndrome for whom thalidomide treatment induced rapid clinical remission and permitted complete corticosteroid withdrawal without clinical relapse.

HIV-1 reservoirs in breast milk and challenges to elimination of breast-feeding transmission of HIV-1.

By compensating for the relative immaturity of the neonatal immune system, breast milk and breast-feeding prevent deaths in children. Nevertheless, transmission of HIV-1 through breast-feeding is responsible for more than half of new pediatric HIV infections. Recent studies of possible HIV-1 reservoirs in breast milk shed new light on features that influence HIV-1 transmission through breast-feeding. The particular characteristics of breast milk CD4(+) T cells that distinguish them from circulating blood lymphocytes (high frequency of cell activation and expression of memory and mucosal homing markers) facilitate the establishment of HIV-1 replication. Breast milk also contains a plethora of factors with anti-infectious, immunomodulatory, or anti-inflammatory properties that can regulate both viral replication and infant susceptibility. In addition, CD8(+) T lymphocytes, macrophages, and epithelial cells in breast milk can alter the dynamics of HIV-1 transmission. Even during efficient antiretroviral therapy, a residual stable, CD4(+) T cell-associated reservoir of HIV-1 is persistently present in breast milk, a likely source of infection. Only prophylactic treatment in infants--ideally with a long-acting drug, administered for the entire duration of breast-feeding--is likely to protect HIV-exposed babies against all forms of HIV transmission from breast milk, including cell-to-cell viral transfer.

Multicytokine detection improves latent tuberculosis diagnosis in health care workers.

In a low-incidence setting, health care workers (HCW) are at a higher risk of tuberculosis than the general population. The suboptimal sensitivity of the QuantiFERON-TB Gold In-Tube (QFT) test remains a critical issue when identifying occupational latent tuberculosis infection (LTBI) in HCW. The aim of this study was to identify additional biomarkers in order to overcome the limits of gamma interferon (IFN-γ) release assays (IGRAs) and improve the performance of LTBI diagnosis within this population. Seventy Bacille Calmette-Guérin-vaccinated HCW regularly exposed to Mycobacterium tuberculosis were grouped according to QFT results into an LTBI-positive group (positive QFT, n = 8), an LTBI-negative group (normal QFT and negative tuberculin skin test [TST], n = 21), and an undetermined group (subpositive QFT and/or positive TST, n = 41). The secretion of 22 cytokines in response to QFT-specific stimulation was quantified using a multiparameter-based immunoassay. As a result, thresholds discriminating LTBI-positive from LTBI-negative HCW were established by comparing areas under the receiver operating characteristic curves for interleukin-2 (IL-2), IL-15, IFN-γ-induced protein 10 (IP-10), and the monokine induced by IFN-γ (MIG), which are biomarkers differentially secreted by the two groups. The combination of IL-15 and MIG provided a sensitivity of 100% and a specificity of 94.1% in distinguishing LTBI-positive from LTBI-negative HCW. When using IL-15 and MIG among the undetermined group, 6/45 HCW could be classified in the LTBI-positive group. The use of additional biomarkers after IGRA screening could improve the diagnosis of LTBI. The performance of these biomarkers and their use in combination with TST and/or QFT, as well as the cost-effectiveness of such a diagnostic strategy, should be evaluated in further larger clinical trials.

Comparison of serum HBsAg quantitation by four immunoassays, and relationships of HBsAg level with HBV replication and HBV genotypes.

BACKGROUND: The decline in hepatitis B virus surface antigen (HBsAg) may be an early predictor of the viral efficacy of Hepatitis B virus (HBV) therapy. The HBsAg levels obtained by different immunoassays now need comparing and the relationships between levels of HBsAg and HBV DNA alongside HBsAg and genotype must be evaluated. METHODOLOGY/PRINCIPAL FINDINGS: HBsAg levels were compared among 80 patients using the Abbott Architect assay, a commercial immunoassay approved for HBsAg detection and quantitation, and three other assays derived from immunoassays approved for HBsAg detection (manufactured by Diasorin, Bio-Rad and Roche). Good correlation was found between the Abbot vs. Diasorin, Bio-Rad and Roche assays with narrow 95% limits of agreement and small mean differences: -0.06 to 0.11, -0.09 log(10) IU/mL; -0.57 to 0.64, -0.04 log(10) IU/mL; -0.09 to 0.45, -0.27 log(10) IU/mL, respectively. These agreements were not affected by genotypes A or D. HBsAg was weakly correlated with HBV DNA, whatever the HBsAg assay used: Abbott, ρ = 0.36 p = 0.001, Diasorin ρ = 0.34, p = 0.002; Bio-Rad ρ = 0.37, p<0.001; or Roche ρ = 0.41, p<0.001. This relationship between levels of HBsAg and HBV DNA seemed to depend on genotypes. Whereas HBsAg (Abbott assay) tended to correlate with HBV DNA for genotype A (ρ = 0.44, p = 0.02), no such correlation was significant for genotypes D (ρ = 0.29, p = 0.15). CONCLUSION/SIGNIFICANCE: The quantitation of HBsAg in routine clinical samples is comparable between the reference assay and the adapted assays with acceptable accuracy limits, low levels of variability and minimum discrepancy. While HBsAg quantitation is not affected by HBV genotype, the observed association between levels of HBsAg and HBV DNA seems genotype dependent.