Canine leishmaniasis: identification of asymptomatic carriers by polymerase chain reaction and immunoblotting.

A major limitation to the study of the epidemiology of canine visceral leishmaniasis is the inability to identify and count asymptomatic carriers because classic diagnositc tests are insufficiently sensitive. We investigated the capacity of the polymerase chain reaction (PCR) to detect the parasite and immunoblotting to detect specific antibodies in samples from dogs living in an endemic area without any symptoms of leishmaniasis. Results of classic serologic tests (immunofluorescence and enzyme-linked immunosorbent assay) were negative. Two independent PCR assays detected the parasite in skin and conjunctival samples from 80% of the dogs. We found specific antibodies by immunoblot in 66% and 56% of the dogs had both specific antibodies and parasite DNA. As controls, samples from dogs with clinical manifestations of the disease before and after treatment were assayed. The frequency of positive PCR samples decreased after treatment, and although antibody levels decreased with cure of the disease, they remained detectable by immunoblot. Results showed that most of the dogs living in an endemic area had been exposed to Leishmania. Both PCR and immunoblot are sensitive enough to detect asyptomatic infection and could be valuable tools for studies monitoring the transmission of the disease and vaccination trials.

Load detection in subjects with sleep-induced upper airway obstruction.

Respiratory flow-resistive load detection in obese patients has been shown to be impaired. We tested the hypothesis that there is no difference in inspiratory flow-resistive load detection measured in nonobese obstructive sleep apnea patients, nonobese snorers, and normal control subjects. Eleven male obstructive sleep apnea patients and seven male snorers were investigated and compared with 10 normal male control subjects. Severely obese patients (body mass index, BMI > 35 kg/m2) were excluded. Patients were investigated by nocturnal polysomnography with measurement of esophageal pressure (Pes). Awake pulmonary function tests were performed before the investigation. Airway resistance (Raw) and lung volumes were measured with plethysmography. Resistive loads were investigated according to Tapper and associates (13) and Killian and associates (12). Resistances were applied for the duration of one inspiratory cycle and a minimum of two breaths allowed between each resistive load. Six different resistances plus background shams were presented 10 times in random order. Flow, pressure, and subject response were recorded with a calibrated multichannel recorder. Subjects signaled detected changes of inspiratory resistance with a hand-held signaling device. The probability of detecting a particular resistance was calculated as the ratio of correct identification to the number of presentations (i.e., 10). The resistance corresponding to a 0.5 probability of detection was determined. The Weber fraction (wf) calculated as delta R/R(apparatus) +Raw. There were no differences between nonobese subjects and controls in terms of resistive load detection.

Comparison of PCR with direct examination of bone marrow aspiration, myeloculture, and serology for diagnosis of visceral Leishmaniasis in immunocompromised patients.

A PCR assay amplifying a repeated sequence from the Leishmania infantum genome was compared with direct examination of bone marrow aspirate, myeloculture, and serology for the diagnosis of visceral leishmaniasis in immunocompromised patients. Of 73 patients living in an area endemic for leishmaniasis and where visceral leishmaniasis was suspected by physicians, only 10 had an indisputable diagnosis of visceral leishmaniasis. None of the diagnostic tests performed in the study achieved 100% sensitivity for diagnosing visceral leishmaniasis. PCR exhibited superior sensitivity (82%) in comparison with bone marrow aspirate examination (55%) and myeloculture (55%). Our PCR assay also showed good specificity (97%), negative predictive value (97%), and positive predictive value (82%) even when all unconfirmed PCR results were scored as false positives. Serology exhibited good sensitivity (80%) and excellent specificity (100%), negative predictive value (98%), and positive predictive value (100%) in diagnosing new cases of visceral leishmaniasis but failed to diagnose relapses. We also observed consistent negative serological results using several different immunological detection methods for 2 of the 10 patients with confirmed cases of visceral leishmaniasis. This lack of serological reactivity persisted throughout the course of their infections. These results demonstrate the importance of using PCR as an aid in the diagnosis of visceral leishmaniasis in immunocompromised patients.

Characterization of a circulating antigen involved in immune complexes in visceral leishmaniasis patients.

Complications of visceral leishmaniasis are explained in part by immune complex pathology, particularly nephritis. Using the polyethylene glycol precipitation method, we identified among these complexes a Leishmania infantum circulating antigen by sodium dodecyl sulfate-polyacrylamide gel electrophoresis and immunoblotting. Monoclonal antibodies were produced to specifically detect this antigen. This protein (molecular weight 51 kD) is a cytoplasmic antigen of L. infantum and its synthesis by promastigotes is related to stationary-phase culture. It was present in sera from dogs whose leishmaniasis was associated with nephritis, and it was found in the sera of 30% of human patients with visceral leishmaniasis. It was also detected in the sera of patients with cutaneous leishmaniasis, and it persists in the sera of clinically cured subjects.

Western blot analysis of antibodies to Leishmania infantum antigens: potential of the 14-kD and 16-kD antigens for diagnosis and epidemiologic purposes.

When infected with Leishmania species, patients develop specific antibodies that constitute the basis of serodiagnosis. Using Western blot analysis, we studied the specificity of anti-L. infantum antibodies in patients with visceral leishmaniasis (including patients with acquired immunodeficiency syndrome [AIDS]) and in healthy subjects living in an endemic area. Sera from patients with visceral leishmaniasis recognized numerous antigens that had a molecular mass range of 12-120 kD. The 14-, 16-, 28-30-, 46-, and 68-kD antigens were recognized by 92%, 95%, 63%, 80%, 69%, and 89% of the patients' sera, respectively. The 14-16-kD antigens had the greatest specificity for leishmaniasis. The same pattern was found with sera from AIDS patients with proven leishmaniasis, but the 14-kD band was not present in some cases; recognition of the 16-kD band was constant. In these patients, Western blotting characterized specific antibodies even when the results of classic serologic tests (indirect immunofluorescent antibody test and enzyme-linked immunosorbent assay) were negative. Western blotting was found to be more sensitive than the IFA and ELISA, and it was used to detect antibodies to the 14-, 16-, 22-, and 24-kD antigens in subjects living in an endemic area. The detection of antibodies for the 14-kD and 16-kD Leishmania antigens would be a valuable tool both in the diagnosis of visceral leishmaniasis and in epidemiologic studies.

Evidence of subjects sensitized to Leishmania infantum on the French Mediterranean coast: differences in gamma interferon production between this population and visceral leishmaniasis patients.

The Marseilles region is an endemic area for visceral mediterranean leishmaniasis, but although the number of dog cases, the parasite's main host, is very high, only a few people develop the disease. We looked for sensitized healthy subjects among 25 healthy individuals living in this area by studying their in vitro lymphoproliferative response to Leishmania infantum antigens and gamma interferon synthesis. We found that 65% of tested subjects were sensitized against L. infantum. We compared their cell mediated immunity to that of 13 active Kala-Azar patients and 13 controls from non-endemic areas. In patients, results showed a specific cellular immuno-deficiency in the lymphocyte response to L. infantum antigens and a global deficiency of gamma interferon production. Interestingly, the healthy individuals from the endemic area who responded to L. infantum antigens were found to produce high gamma interferon levels after L. infantum antigen stimulation. After healing, the cell mediated-immunity of the 3 patients we followed up was similar to that of the sensitized tested healthy subjects, but the former were still producing antibodies at the time of study.

[Study of cellular immunity in patients with Mediterranean visceral leishmaniasis]. / Etude de l'immunité cellulaire chez des malades atteints de leishmaniose viscérale méditerranéenne.

Cellular immunity against Leishmania infantum antigens was studied in visceral leishmaniasis patients and healthy subjects living in a endemic area. Only the healthy subjects were TTL positive with production of gamma interferon, whereas the visceral leishmaniasis patients presented a transitory inhibition of their specific cellular response mechanisms.

Vaccination trial against canine visceral leishmaniasis. Phocean Veterinary Study Group on Visceral Leishmaniasis.

In a double-blind study 393 seronegative dogs, residing in a holoendemic area for Leishmania donovani infantum infection, were randomly assigned to an immunization with a partly purified L.d. infantum-derived preparation, or received adjuvant only. During the first year of the study period the rate of infection was significantly higher in the vaccinated group than in the control one (P less than 0.05), but this difference disappeared during the second year (P = 0.44). Since a similar immunization protocol conferred resistance against experimental murine leishmaniasis, these results stress the differences that may exist between the natural hosts of Leishmania parasites and experimental animal substitutes.

Mediterranean spotted fever in the South of France; serosurvey of dogs.

The authors tested 481 dog sera from the South of France for Rickettsia conorii antibodies using an Indirect Immunofluorescent Antibody Technique. Eighty percent of the sera were positive at a dilution of 1:32 and 45% at a dilution of 1:128. The authors conclude that this provides evidence of the endemic situation of Mediterranean Spotted Fever in the South of France.

[Persistence of visceral leishmaniasis in the southeast of France and recorded frequency of the disease in adults. Apropos of recent cases]. / Persistance de la leishmaniose viscérale dans le sud-est de la France et fréquence marquée de l'affection a l'age adulte. A propos d'observations récentes.

The authors report on 9 cases of mediterranean visceral leishmaniasis (Kala-Azar) collected in the south-eastern part of France in 1978. They emphasize the more frequent occurrence of this disease in adults than children. Out of the 40 cases detected in this region between January 1975 and December 1978, 22 affected adults (55 percent) moreover, Kala-Azar appears to occur particularly in adults with decreased general resistance and diagnosis may be difficult because classical clinical features are no complete.

[Results of a year's mycologic and immunologic surveillance of Candida in an intensive care ward]. / Bilan d'une année de surveillance mycologique et immunologique des Candida dans un service de réanimation.

Patients hospitalized in an intensive care unit must be considered to be exposed to a "high risk" in terms of candida infection. They often combine several of the factors favouring the development of "candida disease": generally deficient state, multiple antibiotics, iatrogenic or pathological immune depression. While certain patients who are admitted are already suffering from "candida infestation" (endogenous localization), the possibilities of exagenous infection are numerous. This study involved 63 patients spending at least one week in the department, from September 1977 to September 1978. Mycological (pharyngeal swab, urine culture, blood culture) and immunological (hemagglutination, immunoelectrophoresis, immunofluorescence) studies were carried out routinely on admission and then every week. Amongst the 63 patients studied, 12 were admitted with negative serology, which became positive during the first 10 days. 5 already had positive serology at the time of admission. From a mycological standpoint, the pharyngeal swab was frequently positive on admission (17 times out of 63). In 63 patients, the infection was acquired during the course of the hospital stay and, in this case, positive urine cultures were more regularly associated with a positive pharyngeal culture.

[Overview of dermatophytes in Mali]. / Panorama des dermatophytes du Mali.

517 isolations of dermatophytes collected in Mali are reported. The various species are described and the main findings are: --Trichophyton species are more common (85.5% isolated from scalp lesions and 77.5% from skin scrapings) than Microsporum species. --Trichophyton verrucosum and Microsporum canis have been found found for the first time in Mali. --Ill-defined clinical forms of scalp lesions caused by either T. schonleinii or T. soudanense occur in the subsaharian areas of the country.

[Biological diagnosis of leishmaniasis (author's transl)]. / Le diagnostic biologique des leishmanioses.

Serochematological, immunological and direct parasitological methods used in the diagnosis of man, dog and wild host reservoirs of Leishmaniasis are described. From their own investigations of more than 5.000 cases, the authors compare the respective technical advantages and the specific indications of each method.

[Leishmaniasis in south-eastern France- -ecology, epidemiology, prophylaxis]. / Les leishmanioses du sud-est de la France ecologie - epidemiologie - prophylaxie

A study of the ecology, epidemiology and prophylaxis of Leishmaniasis in the southeast of France is reported; different methods of the survey are recorded. Between 1965 and 1975, 972 cases of canine Leishmaniasis and from 1968 to 1975 89 cases of visceral human Leishmaniasis and only 3 cases of oriental sore were observed in the "Bouches du Rhône", "Var" and "Vaucluse" Departments. The strains isolated from canine and human visceral Leishmaniasis had a malate dehydrogenase XI whereas Leishmania tropica had a MDH I; no wild animals have been found with Leishmaniasis. The suburbs of the towns, the hills in the center of Marseilles or surrounding Toulon as well as the villages are the principal foci of Phlebotomus perniciousus (96%) as the vector. The biotopes are isolated houses with little gardens surrounded by dry-stone walls exhibiting holes named "barbacanes", with chalky soil and xerophytes. The killing of infected dogs is the best protection of men whereas keeping dogs in sandfly-proof kennels from twilight until one hour after sunrise is best in order to protect this animal. This is quite different from the observations made by RIOUX in the Cevennes.