Minimal intervention dentistry II: part 1. Contribution of the operating microscope to dentistry.

The different aspects of treatment of periodontal disease and mucogingival defects all require an accurate diagnosis in addition to good control and precision during therapeutic procedures. Magnification aids and microsurgery, combined with minimally invasive techniques, can best meet these requirements. The suitability of treatment, the healing time, pain levels and postoperative scarring are all improved and the patient benefits.

[Filovirus]. / Les filovirus.

Since forty years Marburg and Ebola viruses emerge frequently in Africa and are responsible of viral hemorragic fever outbreaks with high mortality rate. Despite intensive research programs, these viruses remain mysterious: the reservoir is not clearly defined, and the mechanisms leading to their high pathogenicity are poorly understood; a defective or inadapted immune response seems to be the main factor. No specific treatment nor vaccine are available for humans. But encouraging results have been obtained in the treatment of filovirus infections in non human primate model with different products, as recombinant nematode anticoagulant protein, anti sens phosphorodiamidate morpholino oligomers or small interfering RNA.As vaccines, recombinantVSV expressing the GP of filovirus or adenovirus expressing the GP and NP of filovirus are very promising in macaque models.

[Monkey-pox, a model of emergent then reemergent disease]. / Le monkey-pox, un paradigme de maladie émergente puis réémergente.

The recent emergence of monkey pox in the United States of America highlights the problem (known for other infectious agents) of dissemination of pathogens outside their endemic area, and of subsequent global threats of variable gravity according to agents. It is a real emergency since monkey pox had been confined to Africa for several decades, where small epidemics occurred from time to time, monkey pox is a "miniature smallpox" which, in Africa, evolves on an endemic (zoonotic) mode with, as reservoirs, several species of wild rodents (mainly squirrels) and some monkey species. It can be accidentally transmitted to man then develops as epidemics, sometimes leading to death. The virus was imported in 2003 in the United States of America, via Gambia rats and wild squirrels (all African species), and infected prairie dogs (which are now in fashion as pets), then crossed the species barrier to man. In the United States of America, screening campaigns, epidemiological investigations, and subsequent treatments led to a rapid control of the epidemic, which is a model of emergent disease for this country. Therapeutic and preventive measures directly applicable to monkey pox are discussed. They can also be applied against other pox virus infections (including smallpox). The risk of criminal introduction of pox viruses is discussed since it is, more than ever, a real worldwide threat.

Inflammatory responses in Ebola virus-infected patients.

Ebola virus subtype Zaire (Ebo-Z) induces acute haemorrhagic fever and a 60-80% mortality rate in humans. Inflammatory responses were monitored in victims and survivors of Ebo-Z haemorrhagic fever during two recent outbreaks in Gabon. Survivors were characterized by a transient release in plasma of interleukin-1beta (IL-1beta), IL-6, tumour necrosis factor-alpha (TNFalpha), macrophage inflammatory protein-1alpha (MIP-1alpha) and MIP-1beta early in the disease, followed by circulation of IL-1 receptor antagonist (IL-1RA) and soluble receptors for TNFalpha (sTNF-R) and IL-6 (sIL-6R) towards the end of the symptomatic phase and after recovery. Fatal infection was associated with moderate levels of TNFalpha and IL-6, and high levels of IL-10, IL-1RA and sTNF-R, in the days before death, while IL-1beta was not detected and MIP-1alpha and MIP-1beta concentrations were similar to those of endemic controls. Simultaneous massive activation of monocytes/macrophages, the main target of Ebo-Z, was suggested in fatal infection by elevated neopterin levels. Thus, presence of IL-1beta and of elevated concentrations of IL-6 in plasma during the symptomatic phase can be used as markers of non-fatal infection, while release of IL-10 and of high levels of neopterin and IL-1RA in plasma as soon as a few days after the disease onset is indicative of a fatal outcome. In conclusion, recovery from Ebo-Z infection is associated with early and well-regulated inflammatory responses, which may be crucial in controlling viral replication and inducing specific immunity. In contrast, defective inflammatory responses and massive monocyte/macrophage activation were associated with fatal outcome.

Human asymptomatic Ebola infection and strong inflammatory response.

BACKGROUND: Ebola virus is one of the most virulent pathogens, killing a very high proportion of patients within 5-7 days. Two outbreaks of fulminating haemorrhagic fever occurred in northern Gabon in 1996, with a 70% case-fatality rate. During both outbreaks we identified some individuals in direct contact with sick patients who never developed symptoms. We aimed to determine whether these individuals were indeed infected with Ebola virus, and how they maintained asymptomatic status. METHODS: Blood was collected from 24 close contacts of symptomatic patients. These asymptomatic individuals were sampled 2, 3, or 4 times during a 1-month period after the first exposure to symptomatic patients. Serum samples were analysed for the presence of Ebola antigens, virus-specific IgM and IgG (by ELISA and western blot), and different cytokines and chemokines. RNA was extracted from peripheral blood mononuclear cells, and reverse transcriptase-PCR assays were done to amplify RNA of Ebola virus. PCR products were then sequenced. FINDINGS: 11 of 24 asymptomatic individuals developed both IgM and IgG responses to Ebola antigens, indicating viral infection. Western-blot analysis showed that IgG responses were directed to nucleoprotein and viral protein of 40 kDa. The glycoprotein and viral protein of 24 kDa genes showed no nucleotide differences between symptomatic and asymptomatic individuals. Asymptomatic individuals had a strong inflammatory response characterised by high circulating concentrations of cytokines and chemokines. INTERPRETATION: This study showed that asymptomatic, replicative Ebola infection can and does occur in human beings. The lack of genetic differences between symptomatic and asymptomatic individuals suggest that asymptomatic Ebola infection did not result from viral mutations. Elucidation of the factors related to the genesis of the strong inflammatory response occurring early during the infectious process in these asymptomatic individuals could increase our understanding of the disease.

Cellular responses to Loa loa experimental infection in mandrills (Mandrillus sphinx) vaccinated with irradiated infective larvae.

In order to shed light on the mechanisms of antifilarial protective immunity, we investigated the course of experimental loaiosis after vaccination in a nonhuman primate host, Mandrillus sphinx. Six vaccinated (V) mandrills received 50 irradiated L3 while six nonvaccinated (NV) received saline solution on days -60, -30 and -15. All animals were challenged with 100 intact L3 (day 0). Parasitological and immunological status were followed for 9 months. Vaccination delayed the appearance and mean peak of microfilaraemia. Five mandrills (Mf-) were never microfilaraemic (one V mandrill) or microfilaraemic on only one occasion (2 V and 2 NV), the other seven having stable microfilaraemia (Mf+). The cytokine response of peripheral blood mononuclear cells to L3 (L3 Ag) was Th2 dominated, while microfilariae (Mf Ag) elicited a Th0-like response. During vaccination, Th2 cytokine production significantly increased in V mandrills against L3 Ag, as well as Mf Ag, whereas Th1 cytokines decreased. On day 60 postinoculation, cellular proliferation was higher in V mandrills in response to L3 and Mf Ags and PHA-L mitogen. At the end of prepatency (on day 130), mandrills with delayed appearance of microfilaraemia exhibited a high, transient IL-2 and IL-4 secretion in response to L3 Ag. Finally, high anti-Mf Th2 cytokine levels characterized Mf-mandrills not only during prepatency, but also (for IL-5) before immunization. However, the presence of a balanced Th1 anti-L3 response during prepatency in the amicrofilaraemic mandrill suggests its importance in protective immunity. Taken together, our data suggest that Th2 cells and also Th1 components of the antifilarial response, especially to larval antigen, may contribute to parasite elimination.

Diagnosis of Ebola haemorrhagic fever by RT-PCR in an epidemic setting.

This study reports the first field evaluation of a new diagnostic technique for Ebola virus disease with sensitivity and specificity. Ebola virus causes rare but fulminating outbreaks in Equatorial Africa. Rapid differentiation from other infections is critical for timely implementation of public health measures. Patients usually die before developing antibodies, necessitating rapid virus detection. A reverse transcriptase-polymerase chain reaction (RT-PCR) assay was developed, implemented and evaluated at Centre International de Recherches Médicales de Franceville (CIRMF) in Gabon, to detect Ebola viral RNA in peripheral blood mononuclear cells (PBMC). Twenty-six laboratory-confirmed patients during and 5 after the acute phase of Ebola haemorrhagic fever, 15 healthy controls and 20 febrile patients not infected with Ebola virus were studied. RT-PCR results were compared with ELISA antigen capture, and Ebola specific IgM and IgG antibody detection. Ebola virus RNA was amplified from 26/26 specimens from the acute phase, 3/5 during recovery, 0/20 febrile patients and 1/15 negative controls. Sensitivity of RT-PCR in identifying acute infection and early convalescence compared with antigen or IgM detection was 100% and 91% respectively, and specificity compared with antigen detection and IgM assay combined was 97%. Antigen capture detected only 83% of those identified by PCR, and IgM only 67%. Ebola virus RNA was detected in all 13 fatalities, only 5 of whom had IgM and none IgG. RT-PCR detected Ebola RNA in PBMC one to three weeks after disappearance of symptoms when antigen was undetectable. RT-PCR was the most sensitive method and able to detect virus from early acute disease throughout early recovery.

HLA class II polymorphism in a Gabonese Banzabi population.

The HLA class II typing of 167 unrelated Gabonese individuals from the Banzabi ethnic group was assessed by polymerase chain reaction-restriction fragment length polymorphism (PCR-RFLP). The most frequent alleles at each locus were DRB1*1501-3 (0.31), DQA1*0102 (0.50), DQB1*0602 (0.42) and DPB1*0402 (0.29). The estimation of the haplotype frequencies as well as the observation of the segregation of several haplotypes using additional HLA typing of relatives, revealed that the three-locus haplotype DRB1*1501-3-DQA1*0102-DQB1*0602 was found at the highest frequency (0.31) among these individuals. This haplotype is not typically African and has already been described in Caucasians, but its presence at high frequency is exclusive to populations originating from Central Africa, and can thus be designated as a particular genetic marker of these populations.

Malaria transmission in a region of savanna-forest mosaic, Haut-Ogooué, Gabon.

During the 2 years 1993 to 1995, an entomological survey was carried out in the savanna-forest area of Franceville, Gabon, investigating malaria transmission in one suburban district of Franceville (Akou) and in one rural village (Benguia). The biting rates of the Anopheles vectors were 10 times higher in the rural zone compared to the suburban zone. Anopheles funestus Giles was the predominant species in both zones followed by Anopheles gambiae s.l. Giles. The densities of Anopheles nili Theobald and Anopheles moucheti Evans were very low. In the suburban zone, transmission was maintained throughout the year by An. funestus and An. gambiae s.l., whereas in rural zones the secondary vectors An. nili and An. moucheti were also involved in transmission. Humans in a suburban setting received one infective bite per person every 4 days, whereas in the rural area the infective biting rate was 4 times higher. Considering each vector, the observed entomological inoculation rates (EIRs) were one infective bite per person every 6 and 17 days for An. funestus and An. gambiae s.l., respectively, at Akou. At Benguia, the EIRs were one infective bite per person every 2, 3, 6, and 19 days for the 4 An. funestus, An. gambiae s.l., An. nili, and An. moucheti, respectively. The predominance of An. funestus over An. gambiae s.l. and its high EIR make it the most important malaria vector in this region of Haut-Ogooué.

Defective humoral responses and extensive intravascular apoptosis are associated with fatal outcome in Ebola virus-infected patients.

Ebola virus is very pathogenic in humans. It induces an acute hemorrhagic fever that leads to death in about 70% of patients. We compared the immune responses of patients who died from Ebola virus disease with those who survived during two large outbreaks in 1996 in Gabon. In survivors, early and increasing levels of IgG, directed mainly against the nucleoprotein and the 40-kDa viral protein, were followed by clearance of circulating viral antigen and activation of cytotoxic T cells, which was indicated by the upregulation of FasL, perforin, CD28 and gamma interferon mRNA in peripheral blood mononuclear cells. In contrast, fatal infection was characterized by impaired humoral responses, with absent specific IgG and barely detectable IgM. Early activation of T cells, indicated by mRNA patterns in peripheral blood mononuclear cells and considerable release of gamma interferon in plasma, was followed in the days preceding death by the disappearance of T cell-related mRNA (including CD3 and CD8). DNA fragmentation in blood leukocytes and release of 41/7 nuclear matrix protein in plasma indicated that massive intravascular apoptosis proceeded relentlessly during the last 5 days of life. Thus, events very early in Ebola virus infection determine the control of viral replication and recovery or catastrophic illness and death.

Ebola hemorrhagic fever outbreaks in Gabon, 1994-1997: epidemiologic and health control issues.

From the end of 1994 to the beginning of 1995, 49 patients with hemorrhagic symptoms were hospitalized in the Makokou General Hospital in northeastern Gabon. Yellow fever (YF) virus was first diagnosed in serum by use of polymerase chain reaction followed by blotting, and a vaccination campaign was immediately instituted. The epidemic, known as the fall 1994 epidemic, ended 6 weeks later. However, some aspects of this epidemic were atypical of YF infection, so a retrospective check for other etiologic agents was undertaken. Ebola (EBO) virus was found to be present concomitantly with YF virus in the epidemic. Two other epidemics (spring and fall 1996) occurred in the same province. GP and L genes of EBO virus isolates from all three epidemics were partially sequenced, which showed a difference of <0.1% in the base pairs. Sequencing also showed that all isolates were very similar to subtype Zaire EBO virus isolates from the Democratic Republic of the Congo.

Plasmodium coatneyi: differential clinical and immune responses of two populations of Macaca fascicularis from different origins.

Two species of macaques, including two Macaca fascicularis from the Philippines, two M. fascicularis from Mauritius, and one Macaca mulatta, were experimentally infected with blood stages of Plasmodium coatneyi and followed during their clinical, parasitological, biological, and immunological evolution. Plasma cytokine (TNF-alpha, IL-1beta, IL-6, IFN-gamma) production peaked for all monkeys 11 days after inoculation, concomitantly with peaks of parasitemia. Only the M. fascicularis from the Philippines survived the infection. The main features, which discriminated nonfatal from fatal cases, were the observation in M. fascicularis from the Philippines of a mean CD4+/CD8+ ratio below I and of their ability, as revealed by mitogenic stimulation of whole blood, to produce increasing amounts of IFN-gamma as infection evolved. The contribution of environmental and genetic factors, which may differentiate the three groups of monkeys and therefore explain fatal or nonfatal evolution of the infection among them, is discussed.

Delta opioid receptors and low temperature myocardial protection.

BACKGROUND: Cardiac surgery continues to be limited by an inability to achieve complete myocardial protection. This may result from the use of hypothermic cardioplegia. Interestingly, the subcellular changes of animal hibernation parallel the altered biology of induced hypothermic myocardial ischemia, but are well tolerated by hibernated mammalian myocardium. Evidence indicates this protection is mediated by activation of the delta opioid receptor, which elicits profound metabolic effects at the whole animal, organ, and cell level. In this study, we sought to determine if pentazocine, with agonist activity at the delta opioid receptor, could improve myocardial recovery following global ischemia over a wide range of temperatures. METHODS: Isolated rabbit hearts received either standard cardioplegia or were pretreated with racemic, d or 1 isomer pentazocine. Hearts were then subjected to 2 hours at 34 degrees C, or 3.5 hours at 20 degrees C, or 4 hours at 10 degrees C of cardioplegic ischemia and reperfused. Functional recovery was compared to controls. RESULTS: Isovolumic developed pressure, coronary flow, oxygen consumption, and ultrastructural preservation were enhanced with pentazocine delta opioid mediated protection, which appears to be additive to standard cardioplegia, even at low temperatures. CONCLUSIONS: Teleologically, delta opioid protection parallels animal hibernation, which occurs from 34 degrees down to 0 degrees C. The use of delta opioid receptor agonists may have important clinical implications for cardiac surgery and deserves further study.

[HTLV-1 infection in time and space]. / HTLV-1 dans le temps et dans l'espace.

Human T-cell leukemia virus type 1 (HTLV-1) is a member of the Oncoretrovirinae family containing several viruses that have been associated with a low incidence of leukemia and sarcoma in mammals. Primates are susceptible to viruses of genera HTLV (humans) and STLV (other primates). The high degree of homology in genomic arrangement of HTLV and STLV is probably due to the existence of a common simian ancestor. Most infections are asymptomatic but a few cases exhibit blood diseases, e.g., T-lymphoma or T-cell leukemia, or neurologic disease, mainly, HTLV-1 associated myelopathy and tropical spastic paraparesia (HAM-TSP). The four major modes of viral transmission are vertical transmission from mother to child either in utero or, more commonly, during breastfeeding, sexual intercourse, blood transfusion, and intravenous drug use. Geographic distribution of HTLV-1 and its confinement to a few well-defined ecosystems have yet to be explained. Diagnosis is now easy and can reduce transmission by intravenous drugs use. Development of a vaccine seems possible given the low genetic variability of this virus.

[Biohazards due to Orthopoxvirus: should we re-vaccinate against smallpox?]. / Les biorisques liés aux Orthopoxvirus: faut-il revacciner contre la variole?

Although the WHO declared global smallpox eradication in 1980, the Orthopoxvirus remains a source of concern for several reasons. Firstly, stocks of the smallpox virus have been preserved for experimental use (at least officially in the USA and Russia) so that an escaped isolate could lead to reemergence and spread of the disease worldwide. Secondly discontinuation of smallpox vaccination programs has led to dwindling acquired immunity in the world population thus raising the risk of epidemic extension of several Orthopoxvirus zoonoses (e.g., monkeypox). Thirdly stocks of camelpox virus which is very similar to Smallpox virus and was intended for biological warfare were discovered during the Gulf War in 1991 and pose a potentially serious threat. Finally official stocks of Smallpox virus could be stolen and used by bioterrorists. Thus reemergence of Orthopoxvirus including smallpox, monkeypox, cowpox, and camelpox is a real danger and contingency planning is needed to define prophylactic and therapeutic strategies to prevent and/or stop an epidemics. Adverse effects from earlier smallpox vaccine (vaccinia) in healthy people or immunocompromised people (congenital or acquired as in HIV infected patients) are absolute contraindications to smallpox vaccination at this time. Further research is needed to develop new vaccines (e.g., attenuated isolates of vaccinia) and effective treatment. This is the only valid reasons for postponing planned destruction of remaining Smallpox virus stocks.

[Ebola virus: what the practitioner needs to know]. / Virus Ebola: l'essentiel pour le praticien.

The Ebola virus is an RNA virus of Filoviridae family. The earliest documented fatal epidemic of Ebola hemorrhagic occurred in 1976. There are four genetically different subtypes of Ebola virus. The virus remains in the blood for several weeks, can maintain its infectivity for several weeks at 20 degrees C outside the body, and survives for several weeks in corpses. Isolation of Ebola virus requires level 4 laboratory security conditions. Specimens are obtained by culturing mammal cells. Identification is achieved using reference serums. Serologic diagnosis is made using mainly ELISA technique for immunocapture of IgM or EBO Ag. The natural reservoir for Ebola virus is unknown. One possibility is that each isolated strain has a different reservoir. In recorded outbreaks, the index case has often had a history of contact with non-human primates. However since these animals are also highly sensitive to the virus, they cannot be considered as reservoirs but only as intermediate hosts. Transmission requires close contact such as occurs in association with health care, local customs, or funeral rites. In humans, infection causes hemorrhagic fever that progresses to diarrhea within 5 to 10 days. Recovery is observed in only 25% of cases. During outbreaks containment depends on implementation of simple precautions including isolation of suspected cases, appropriate protective clothing, disinfection with hypochlorite solutions, and proper waste disposal.