In eight of the 12 countries tested, antibodies to group O viruses were identified. Numbers of HIV-1 group O viruses are low. Their presence is not restricted to Cameroon and neighbouring countries but can also be found in west and south-east Africa.

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Our results demonstrate that, from Madagascar to Senegal, the epidemiologic and virologic characteristics of influenza viruses are diverse in terms of spatiotemporal circulation of the different virus types, subtypes, and strains. Our data highlight the importance of country-specific surveillance and of data and virus sharing, and they provide a rational basis to aid policy makers to develop strategies, such as vaccination at the right moment and with the right formulation, aimed at reducing the disease burden in Africa and Madagascar.

Pathogens often encounter stressful conditions inside their hosts. In the attempt to characterize the stress response in Brucella suis, a gene highly homologous to Escherichia coli clpB was isolated from Brucella suis, and the deduced amino acid sequence showed features typical of the ClpB ATPase family of stress response proteins. Under high-temperature stress conditions, ClpB of B. suis was induced, and an isogenic B. suis clpB mutant showed increased sensitivity to high temperature, but also to ethanol stress and acid pH. The effects were reversible by complementation. Simultaneous inactivation of clpA and clpB resulted in a mutant that was sensitive to oxidative stress. In B. suis expressing gfp, ClpA but not ClpB participated in degradation of the green fluorescent protein at 42°C. We concluded that ClpB was responsible for tolerance to several stresses and that the lethality caused by harsh environmental conditions may have similar molecular origins.Brucella suis is a gram-negative, facultative intracellular bacterium and one of the causative agents of brucellosis in humans and animals (2). Oral infection, which is considered a natural route of infection by brucellae, and localization of the bacteria inside the host cell phagosome most likely expose the pathogen to various stresses. In response to environmental stress conditions such as elevated temperatures, variation in pH, and starvation, a certain number of proteins are induced or repressed in Brucella spp., among which are the GroE and DnaK heat shock proteins (12,16,24). In intracellular brucellae, which encounter stressful conditions such as acidic pH (23) and possibly oxidative stress due to the presence of oxygen radicals, the known stress proteins GroEL and DnaK are induced during infection, and DnaK is essential for multiplication of B. suis in macrophage-like cells (12,17). The HtrA (high-temperature requirement) stress protein is involved in high-level spleen colonization with Brucella abortus in mice (5). Several other stress response proteins including ClpA, ClpB, ClpC, and ClpX, are members of a family of proteins called the Clp ATPases (HSP100 proteins), represented in prokaryotes and eukaryotes with a high degree of conservation (28). Clp stress proteins can be induced by high temperature, oxidative stress, high salt or ethanol concentration, and iron limitation (14,26,28). In addition to being sensitive to various stresses, Listeria monocytogenes clpC mutants are attenuated for virulence in mice (26).Our article describes the isolation and characterization of a B. suis gene encoding a homolog of the ClpB stress response proteins. An isogenic clpB mutant and the complemented strain were constructed and compared with the wild type and a clpA mutant (4) for survival at high temperature, at different ethanol concentrations, at acid pH, and in the presence of hydrogen peroxide. In addition, the effect of hydrogen peroxide on a clpAB double mutant was studied, and the possible participation of ClpA and ClpB in protein degradation was analyzed....

Background  Many countries in Africa have lacked sentinel surveillance systems for influenza and are under‐represented in data used for global vaccine strain selection. Objectives  We describe 8 years of sentinel surveillance data and the contribution of influenza and other viruses to medically attended influenza‐like illness (ILI) in Côte d’Ivoire. Methods  Sentinel surveillance was established in 2003. Nasopharyngeal (NP) specimens and epidemiologic data are collected from persons of all ages presenting with ILI at sentinel sites. Respiratory specimens have been tested for influenza using various viral and molecular diagnostic methods. A subset of 470 specimens collected from children aged 0–5 years were tested for multiple respiratory viruses using RT‐PCR. Results  From 2003 to 2010, 5074 NP specimens were collected from patients with ILI. Overall, 969/5074 (19%) of these specimens tested positive for influenza. Seasonal influenza A(H1N1) viruses predominated during 5 years and influenza A(H3N2) viruses predominated during 3 years. Influenza B viruses cocirculated with influenza A viruses during each year from 2004 to 2010. Seasonal peaks in influenza circulation were observed during the months of May, June, and October, with the largest peak corresponding with the primary rainfall season. Of 470 specimens collected from children under aged 5 who were tested for multiple respiratory viruses, a viral respiratory pathogen was detected in 401/470 (85%) of specimens. Commonly detected viruses were RSV (113 of 470 specimens, 24%), rhinoviruses (85/470, 18%), influenza (77/470, 16%), and parainfluenza (75/470, 16%). Conclusion  In Côte d’Ivoire, there is a significant annual contribution of influenza and other respiratory viruses to medically attended ILI.

The protein ClpA belongs to a diverse group of polypeptides named ClpATPases, which are highly conserved, and which include several molecular chaperones. In this study the gene encoding the 91 kDa protein b-ClpA of the facultative intracellular pathogen Brucella suis, which showed 70 % identity to ClpA of Rhodobacter blasticus, was identified and sequenced. Following heterologous expression in Escherichia coli strains SG1126 (∆clpA) and SG1127 (∆lon ∆clpA), b-ClpA replaced the function of E. coli ClpA, participating in the degradation of abnormal proteins. A b-clpA null mutant of B. suis was constructed, and growth experiments at 37 and 42 SC showed reduced growth rates for the null mutant, especially at the elevated temperature. The mutant complemented by b-clpA and overexpressing the gene was even more impaired at 37 and 42 SC. In intracellular infection of human THP-1 or murine J774 macrophage-like cells, the clpA null mutant and, to a lesser extent, the strain of B. suis overexpressing b-clpA behaved similarly to the wild-type strain. In a murine model of infection, however, the absence of ClpA significantly increased persistence of B. suis. These results showed that in B. suis the highly conserved protein ClpA by itself was dispensable for intramacrophagic growth, but was involved in temperature-dependent growth regulation, and in bacterial clearance from infected BALB/c mice.

The heat shock protein DnaK is essential for intramacrophagic replication of Brucella suis. The replacement of the stress-inducible, native dnaK promoter of B. suis by the promoter of the constitutively expressed bla gene resulted in temperature-independent synthesis of DnaK. In contrast to a dnaK null mutant, this strain grew at 37°C, with a thermal cutoff at 39°C. However, the constitutive dnaK mutant, which showed high sensitivity to H 2 O 2 -mediated stress, failed to multiply in murine macrophage-like cells and was rapidly eliminated in a mouse model of infection, adding strong arguments to our hypothesis that stress-mediated and heat shock promoter-dependent induction of dnaK is a crucial event in the intracellular replication of B. suis.We have described the important role of the heat shock protein and molecular chaperone DnaK in intramacrophagic growth of Brucella suis and its acid-induced expression (12). Brucella spp., facultative intracellular, gram-negative bacteria which are the causative agents of brucellosis in humans and animals (6), resist intracellular killing and replicate within the phagosome of macrophages (3,11,16). It has been observed recently that this phagosome is acidic (pH 4.0 to 4.5), suggesting a stressful environment, and that early acidification is essential for intracellular multiplication of B. suis (19). Work on the identification of proteins induced under stress conditions in Brucella abortus and Brucella melitensis confirmed our results (17,20,23). From previous data (12), we concluded that DnaK from B. suis may play an essential role as part of protein repair systems in protecting the bacteria from the environment encountered in the phagosome. Another hypothesis is that DnaK may be directly involved in folding and proper localization of virulence factors, as intracellular multiplication is abolished in the null mutant.The earlier work was done with a dnaK null mutant, allowing us to conclude only that the chaperone participated in intracellular multiplication of B. suis (12). On the other hand, dnaJ, which is located downstream of dnaK and forms an operon with the latter, is not involved in resistance to acid stress and intracellular multiplication of the pathogen (12). As a dnaJ knockout mutant of B. suis behaves like the wild-type strain with respect to these properties, we concluded that dnaJ was of no relevance in the context of the work presented here. Western blot analysis showed induction of dnaK during heat and acid shock and under intramacrophagic growth conditions (12). We therefore hypothesized that low-level, constitutive expression of dnaK was not sufficient for B. suis to resist macrophage attack. To verify this hypothesis, we replaced the original dnaK promoter on the B. suis chromosome by the constitutive promoter of the ␤-lactamase gene bla TEM (P bla ) from pUC18, and we studied the phenotype of the mutant obtained. P bla was chosen, as we and others (13,14) have observed that bla is expressed in brucellae, conferring ampicillin resistance to strains bearin...

f Tuberculosis remains a major health problem due to the emergence of drug-resistant strains of Mycobacterium tuberculosis. Some models have provided valuable information about drug resistance and efficacy; however, the translation of these results into effective human treatments has mostly proven unsuccessful. In this study, we adapted high-content screening (HCS) technology to investigate the activities of antitubercular compounds in the context of an in vitro granuloma model. We observed significant shifts in the MIC 50 s between the activities of the compounds under extracellular and granuloma conditions. T uberculosis (TB) is a respiratory disease that is one of the major causes of mortality and morbidity worldwide. Almost 20 people develop TB and four patients die from the disease every minute somewhere in the world (1). The disease can be cured by drug treatment involving a regimen of several drugs. The World Health Organization (WHO) estimates that there were around 0.5 million new cases of multidrug-resistant TB (MDR-TB) in 2012 (2, 3). A recent report confirmed that MDR-TB is a continuing worldwide health problem, even in high-income countries with a low incidence of TB (4).The immune system undoubtedly plays a major role in TB control (5). Signaling events of the immune system lead to the formation of a granuloma, the hallmark of TB. Granulomas are an immune microenvironment in which the infection can be controlled but also a niche in which bacilli can thrive and modulate immune responses to ensure their survival for long periods without causing damage (6, 7). Granulomas are cell aggregates that form tridimensional and heterogeneous structures (8, 9). We previously described an in vitro granuloma model involving human blood cells either infected with mycobacteria or incubated with mycobacterial antigens (8, 10). Either of these models resulted in the formation of a typical epithelioid granuloma with morphological characteristics and properties of cellular differentiation similar to those of natural granulomas (8,10,11). This model may help bridge the gap between the in vitro evaluation of MICs and costly in vivo efficacy studies in guinea pigs. In particular, compounds that are effective in the in vitro granuloma model could be prioritized for guinea pig studies.High-content screening (HCS) technology has been used to identify anti-TB compounds (12)(13)(14). This technology has several advantages over traditional phenotypic screening approaches. This technology has mostly been used in single-cell experiments, because it is adapted to identify and analyze images (12, 13); however, in this paper, we focus on the use of granulomas instead of single cells. In this report, we describe the development of a novel HCS method to evaluate the activities of reference compounds against a green fluorescent protein (GFP)-expressing H37Rv strain of Mycobacterium tuberculosis (MTB-GFP) within 5 days, and we compared the MIC 50 values with those of classical liquid cultures (extracellular), intracellular growth (singl...