Survey of laboratory-acquired infections around the world in biosafety level 3 and 4 laboratories.

Laboratory-acquired infections due to a variety of bacteria, viruses, parasites, and fungi have been described over the last century, and laboratory workers are at risk of exposure to these infectious agents. However, reporting laboratory-associated infections has been largely voluntary, and there is no way to determine the real number of people involved or to know the precise risks for workers. In this study, an international survey based on volunteering was conducted in biosafety level 3 and 4 laboratories to determine the number of laboratory-acquired infections and the possible underlying causes of these contaminations. The analysis of the survey reveals that laboratory-acquired infections have been infrequent and even rare in recent years, and human errors represent a very high percentage of the cases. Today, most risks from biological hazards can be reduced through the use of appropriate procedures and techniques, containment devices and facilities, and the training of personnel.

Cardiovascular risk and fitness in veteran football players.

Veteran football players above 40 years have rarely been subject to scientific investigations. This is worrisome because their number is considerable and their cardiovascular risk probably increased. Therefore, a cross-sectional study was conducted in 100 football players between 40 and 63 years of age. This included a medical history and physical examination, venous blood sampling, measurement of resting blood pressure, a resting electrocardiogram (ECG), an exhaustive cycle ergometry and a multistage field test. Also, measurements of heart rate and blood lactate concentration were carried out during one typical training session and one match. Participants trained 1.0 ± 0.6 sessions per week and played 27 ± 8 matches per season. Of them, 19% were smokers. Resting blood pressure was 138 ± 15/88 ± 8 mmHg. Hypertension prevalence (WHO definition) was 66%. Total cholesterol averaged 220 ± 41 mg . dl(-1), HDL 46 ± 13 mg . dl(-1) and LDL 134 ± 33 mg . dl(-1). The average 10-year risk for cardiovascular events (Framingham score) was 6%. Mean maximal power output on the cycle ergometer was 2.8 ± 0.6 W . kg(-1), mean VO2peak 40.0 ± 7.3 ml . min(-1) . kg(-1). Comparing training and competition, no significant differences in cardiovascular and metabolic load were found. In summary, their cardiovascular risk was similar to age-adjusted reference values. However, they showed slightly better ergometric performance. More frequent training stimuli might be necessary to reach more favourable risk factor profiles. Training and competition lead to similar cardiocirculatory and metabolic stress which is considerably high and might put players into danger who have pre-existing cardiac disease.

Emergence of Plasmodium ovale malaria among the French Armed Forces in the Republic of Ivory Coast: 20 years of clinical and biological experience.

BACKGROUND: French military surveillance identified an increase in Plasmodium ovale attacks among soldiers in Ivory Coast. This emergence and the low sensitivity of biological tests raise the question of a possible role of P. ovale variant species. METHODS: Epidemiological data about P. ovale attacks from 1993 to 2012 were studied; the species diagnosis was based on a thin blood smear and/or a quick diagnostic test. Clinical and biological features in soldiers hospitalized in 2 French military hospitals were also reviewed. Malaria polymerase chain reaction followed by genotyping was performed when available. RESULTS: French military physicians declared 328 P. ovale attacks over the 20-year study. A peak of incidence occurred in 2005. Among patients with positive blood smears, the quick diagnostic test was positive in 33 of 101 tests performed. The hospital study showed that symptoms and biological changes were not specific, which made diagnosis challenging: fever, anemia, and thrombocytopenia were not present in 20%, 71%, and 23% of the 45 confirmed cases, respectively. It was possible to perform molecular investigations on 19 clinical isolates: 18 were classic haplotypes with additional polymorphism and 1 was variant. CONCLUSIONS: This emergence of P. ovale malaria enabled a good description to be made in nonimmune patients. The lack of sensitivity of both clinical features and quick diagnostic tests suggests an underestimation. Reasons for this outbreak are especially intense exposure to the vectors and the insufficient efficacy of doxycycline against P. ovale. The polymorphism of classic haplotypes of P. ovale rather than variant forms could be involved.

Sequence specific alkylation of DNA by hairpin pyrrole-imidazole polyamide conjugates.

BACKGROUND: Pyrrole-imidazole polyamides are synthetic ligands that recognize predetermined sequences in the minor groove of DNA with affinities and specificities comparable to those of DNA-binding proteins. As a result of their DNA-binding properties, polyamides could deliver reactive moieties for covalent reaction at specific DNA sequences and thereby inhibit DNA-protein interactions. Site-specific alkylation of DNA could be a useful tool for regulating gene expression. As a minimal first step, we set out to design and synthesize a class of hairpin polyamides equipped with DNA alkylating agents and characterize the specificity and yield of covalent modification. RESULTS: Bis(dichloroethylamino)benzene derivatives of the well-characterized chlorambucil (CHL) were attached to the gamma turn of an eight-ring hairpin polyamide targeted to the HIV-1 promoter. We found that a hairpin polyamide-CHL conjugate binds and selectively alkylates predetermined sites in the HIV promoter at subnanomolar concentrations. Cleavage sites were determined on both strands of a restriction fragment containing the HIV-1 promoter, revealing good specificity and a high yield of alkylation. CONCLUSIONS: The ability of polyamide-CHL conjugates to sequence specifically alkylate double-stranded DNA in high yield and at low concentrations sets the stage for testing their use as regulators of gene expression in cell culture and ultimately in complex organisms.

Fmoc solid phase synthesis of polyamides containing pyrrole and imidazole amino acids.

[structure: see text]. Polyamides containing N-methylimidazole (Im) and N-methylpyrrole (Py) amino acids are synthetic ligands that have an affinity and specificity for DNA comparable to those of many naturally occurring DNA binding proteins. A machine-assisted Fmoc solid phase synthesis of polyamides has been optimized to afford high stepwise coupling yields (>99%). Two monomer building blocks, Fmoc-Py acid and Fmoc-Im acid, were prepared in multigram scale. Cleavage by aminolysis followed by HPLC purification affords up to 200 mg quantities of polyamide with purities and yields greater than or equal to those reported using Boc chemistry. A broader set of reaction conditions will increase the number and complexity of minor groove binding polyamides which may be prepared and help ensure compatibility with many commercially available peptide synthesizers.

Negative impact of laws regarding biosecurity and bioterrorism on real diseases.

Research on highly pathogenic microorganisms in biosafety level 3 and 4 laboratories is very important for human public health, as it provides opportunities for the development of vaccines and novel therapeutics as well as diagnostic methods to prevent epidemics. However, in recent years, after the anthrax and World Trade Center attacks in 2001 in the USA, the threat of bioterrorism has grown for both the public and the authorities. As a result, technical and physical containment measures and biosafety and biosecurity practices have been implemented in laboratories handling these dangerous pathogens. Working with selected biological agents and toxins is now highly regulated, owing to their potential to pose a threat to public health and safety, despite the fact that the anthrax attack was found to be the result of a lack of security at a US Army laboratory. Thus, these added regulations have been associated with a large amount of fruitless investment. Herein, we describe the limitations of research in these facilities, and the multiple consequences of the increased regulations. These limitations have seriously negatively impacted on the number of collaborations, the size of research projects, and, more generally, scientific research on microbial pathogens. Clearly, the actual number of known victims and fatalities caused by the intentional use of microorganisms has been negligible as compared with those caused by naturally acquired human infections.

[Delayed diagnosis of Plasmodium falciparum in a soldier in Uganda: false-positive rapid diagnostic test associated with reduced repeats in pfhrp2]. / Retard de diagnostic d'un accès palustre à Plasmodium falciparum chez un militaire en Ouganda : négativité des tests de diagnostic rapide associée à un polymorphisme de l'antigène HRP2.

Rapid diagnostic tests (RDTs) are the best alternative for malaria diagnosis where a microscopic examination cannot be performed. We report here the first case of P. falciparum (false-negative) misdiagnosis in a soldier stationed in Uganda, associated with a reduced number of repeats in the P. falciparum histidine-rich protein 2 gene (pfhrp2). This gene was subsequently sequenced to determine the reason for the discordance between the RDT results and the later microscopic examination. Ten repeats of the type 2 motif AHHAHHAAD and four repeats of the type 7 motif AHHAAD were found. This isolate belongs to the group of non-sensitive parasites (<43 repeats) that are not detected by HRP2 RDTs. This inappropriate case management could have been fatal for the patient. This case confirms the problem of negative RDT results in isolated situations and of basing a therapeutic strategy on these negative results. Investigations should be conducted in Uganda and other areas of Africa to determine the presence and the geographical spread of parasites with pfhrp2 gene deletion to ensure the best performance of RDTs.

Absence of association between ex vivo susceptibility to doxycycline and pftetQ and pfmdt copy numbers in Plasmodium falciparum isolates from Dakar, Senegal.

The objective of this study was to validate the use of pftetQ and pfmdt genes as molecular markers of decreased in vitro susceptibility to doxycycline in 113 Plasmodium falciparum isolates from Dakar, Senegal. The results show that copy numbers of pftetQ and pfmdt, estimated by TaqMan real-time PCR, are not significantly associated with reduced susceptibility to doxycycline in vitro; however, the number of samples with a high doxycycline IC(50) was likely to be too low to derive statistically significant results. Thus, no definitive conclusions could be drawn. The markers should be further tested by analysing more isolates.

Solid-phase synthesis of DNA binding polyamides on oxime resin.

Control of the energetics and specificity of DNA binding polyamides is necessary for inhibition of protein-DNA complex formation and gene regulation studies. Typically, solid-phase methods using Boc monomers for synthesis have depended on Boc-beta-Ala-PAM resin which affords a beta-alanine-Dp tail at the C-terminus, after cleavage with N,N-dimethylaminopropylamine (Dp). To address the energetic consequences of this tail for DNA minor groove binding, we describe an alternative solid phase method employing the Kaiser oxime resin which allows the synthesis of polyamides with incrementally shortened C-terminal tails. Polyamides without Dp and having methyl amide tails rather than beta-alanine show similar affinity relative to the standard beta-Dp tail. The truncated tail diminishes the A,T base pair energetic preference of the beta-Dp tail which will allow a greater variety of DNA sequences to be targeted by hairpin polyamides.

Inhibition of major groove DNA binding bZIP proteins by positive patch polyamides.

Cell permeable synthetic ligands that bind to predetermined DNA sequences offer a chemical approach to gene regulation, provided inhibition of a broad range of DNA transcription factors can be achieved. DNA minor groove binding polyamides containing aminoalkyl substituents at the N-1 of a single pyrrole residue display inhibitory effects for a bZIP protein which binds exclusively in the DNA major groove. For major groove protein inhibition, specific protein-DNA contacts along the phosphate backbone were targeted with the positively charged dimethylamino substituent on the backbone of a minor groove binding polyamide hairpin. Remarkably, these polyamides bind DNA with enhanced affinity and uncompromised specificity when compared to polyamides with the aminoalkyl moiety at the C-terminus. By adding bZIP transcription factors to the class of protein-DNA complexes that can be disrupted by minor groove binding ligands, these results may increase the functional utility of polyamides as regulators of gene expression.