Larry D. Hendricks

Cutaneous leishmaniasis in Kenya: transmission of Leishmania major to man by the bite of a naturally infected Phlebotomus duboscqi

One leishmanial stock was isolated from a Phlebotomus duboscqi female captured in Baringo District, Kenya, and others from papular lesions that developed at sites where this sandfly had fed on a man. When characterized by cellulose acetate electrophoresis (eight enzymes examined), these isolates proved to be identical to known Leishmania major strains from man and a rodent (Arvicanthis sp.) and different from L. donovani and L. adleri, which also occur in Baringo. This is the first case of human cutaneous leishmaniasis caused by L. major reported from Kenya.

Cutaneous leishmaniasis caused by Leishmania tropica in Kenya

Cutaneous leishmaniasis is endemic in Algeria, but clinical and parasitological data from this area are scarce. In order to document the transmission of this disease in a peri-urban setting, cutaneous lesions from patients living in Constantine City and surrounding areas were spotted on filter paper for diagnosis and species identification using real-time PCR. Surprisingly, Leishmania tropica was detected in 6/69 patients, and confirmation was obtained by sequencing. This observation suggests a modification of the epidemiology of cutaneous leishmaniasis in Algeria and should alert physicians and policy-makers to the risk of antimony treatment failure with this species.9 leishmanial strains, isolated from cutaneous papulonodular lesions on 3 patients, were characterized by cellulose acetate electrophoresis using 7 enzymes. The patterns obtained were indistinguishable from those of a Leishmania tropica reference strain and these 9 strains were similar to L. tropica in failing to infect mice. Although these 3 patients were Americans, their only potential exposure to sandflies was in Kenya, and thus they are believed to be the first cases of cutaneous leishmaniasis due to L. tropica in Kenya.

Leishmania major: the suitability of East African nonhuman primates as animal models for cutaneous leishmaniasis.

The susceptibility of four species of East African nonhuman primates to experimental infection with Leishmania major was investigated. Four Sykes monkeys (Cercopithecus mitis), two vervet monkeys (Cercopithecus aethiops), two baboons (Papio cynocephalus), and two brown bushbabies (Galago garnettii) were each inoculated intradermally on the left eyelid, left ear, and nose with 0.1 ml of medium containing 1 x 10(7) promastigotes of a characterized L. major strain. All the nonhuman primates except the bushbabies developed erythema and conspicuous nodules on the eyelids and ears by 3 weeks PI. The nodules increased rapidly in size and ulceration was evident on the eyelids and ears by 49 days PI in the vervets, Sykes, and baboons. The aspirates were positive in culture or smears at 35, 49, 63, and 77 days PI. No parasites were observed in cultures or smears at 92, 105, 128, 147, and 161 days PI. The lesions in these animals began resolving by 84 days PI and were completely healed by 112 days PI. The exception was one baboon in which lesion healing did not start until around 147 days and was completely healed by 182 days PI. Cultures from the liver failed to demonstrate visceralization of the parasite in any of the animals throughout the 68 weeks of the experiment. Challenge with the same strain of L. major 6 months PI, corresponding to about 3 months after self cure, failed to produce infection in any of these experimental hosts. All the nonhuman primates except the bushbaby when challenged with the same strain of L. major at 12 months PI developed lesions and were positive for parasites at 14 and 28 days PI. Positive cultures were obtained from the eyelid and ear of one vervet up to 42 days PI. However, the lesion sizes in all these animals were smaller than in the initial infection and did not ulcerate. The nodules disappeared within 6 to 8 weeks as compared to 16 weeks in the initial infection. The histopathological appearance of the lesions varied from diffuse infiltration of plasma cells and lymphocytes which increased progressively to granulomata with epitheloid cells. This study shows that the vervets, Sykes, and the baboons are equally susceptible to L. major infection, while bushbabies are refractory. The vervets, Sykes, and baboons demonstrate a self-healing phenomenon within about 3 months which is comparable to that observed in humans infected with L. major. These three species of nonhuman primates are therefore considered as suitable models for drug or vaccine trials against human zoonotic cutaneous leishmaniasis.

Liposomes in leishmaniasis: effects of parasite virulence on treatment of experimental leishmaniasis in hamsters

During studies on the use of liposomes as drug carriers in experimental leishmaniasis in hamsters, we noted incidentally that the apparent virulence of the infection often varied widely between different large groups of animals. When the death rates among control animals (injected only with saline) were compared with hepatic parasite counts of survivors in the same group, three distinctive types of infection were observed: type I, low death rate, low parasite count in survivors; type II, high death rate, low parasite count in survivors; type III, high death rate, high parasite count in survivors. The apparent virulence, based on death rates both at early and late stages of infection, was in the order I less than II less than III. Therapeutic efficacy of a drug (meglumine antimoniate) or liposome-encapsulated drug against each type of infection was in the order I greater than II greater than III. Liposomes reduced the drug dose required for each infection type many hundred-fold and reduced the death rate for type I to zero. However, among animals with type III (or even type II) infection certain individuals were completely refractory to treatment, even when liposome-encapsulated drug was employed, and the lowest mortality rate achieved was approximately 30%. This latter resistance to treatment may have been due to irreversible tissue damage caused by advanced disease, or it may have reflected resistance of certain virulent infections to treatment.

Human cutaneous leishmaniasis caused by Leishmania donovani s.l. in Kenya

Our laboratory is characterizing Leishmania stabilates and isolates from active leishmaniasis cases. Smears and cultures from aspirates made on different dates from a single lesion on the bridge of the nose of an 18 years old Kenyan male from Nyandarua District contained Leishmania. The isolates, NLB-271 and NLB-271-IA, were characterized by cellulose acetate electrophoresis (CAE) using 20 enzyme systems and by Southern analysis using 2 deoxyribonucleic acid (DNA) probes (pDK10 and pDK20) from a Dakar strain of L. major (MHOM/SN/00/DK1) and a third probe, p7-059 from L. infantum strain ITMAP-263. Digestion of the two Leishmania DNAs with endonucleases HindIII and PstI, followed by hybridization with the 3 probes, revealed DNA fragment banding patterns indistinguishable from those of the L. donovani species complex. The CAE isoenzyme profiles of these 2 Kenyan isolates were indistinguishable from those of Kenyan L. donovani strains we designated as zymodeme Z6. Excluding post-kala-azar dermal leishmaniasis, this constitutes the first human case of cutaneous leishmaniasis caused by L. donovani s.l. in Kenya. Previously, cutaneous leishmaniasis cases in Kenya have been due to L. aethiopica, L. major and L. tropica only.

Leishmania mexicana: Purine metabolism in promastigotes, axenic amastigotes, and amastigotes derived from vero cells

Leishmania mexicana mexicana promastigotes, axenic amastigotes, and amastigotes derived from Vero cells were examined for de novo purine synthesis and mechanisms of purine salvage. Both promastigotes and axenic amastigotes were incapable of de novo purine synthesis, as shown by the lack of [14C]formate and [14C]glycine incorporation into purine nucleotide pools. However, the ready incorporation of [14C]hypoxanthine, [14C]adenine, and [14C]guanine suggested that purine salvage pathways were operating. In addition, a significant percentage (greater than or equal to 60%) of the total label from these purine precursors was associated with adenylate nucleotides. Nucleotide pool levels of axenic amastigotes were consistently greater but the specific activities were less than those of promastigotes, suggesting a slower rate of purine metabolism in the axenic amastigote form. Similar results were obtained from amastigotes isolated from infected Vero cells.

The culture of Leishmania donovani in Schneider's insect medium: its value in the diagnosis and management of patients with visceral leishmaniasis

Abstract Schneiders insect culture medium, supplemented by the addition of 30% ( v v ) heat-inactivated foetal bovine serum plus 100 IU penicillin and 100 μm streptomycin (final concentration per ml), is shown to be simple to use in the diagnosis and management of visceral leishmaniasis in Kenya. It is especially valuable because small numbers of parasites can be detected in this culture medium later during the course of treatment.

American cutaneous leishmaniasis: presentation and problems of patient management

We report our experience with the diagnosis and treatment of 60 patients with American cutaneous leishmaniasis. They were infected in Panama (55), Brazil (4) or Colombia (I). Among 35 patients with a 3 week exposure in Panama, the mean maximum incubation period was 33 days (range 4-81 days). Diagnosis was delayed an average of 93 days after onset of skin lesions, due to the patients delay in seeking medical attention (31 days), medical personnels delay in considering the diagnosis (45 days), and the laboratorys delay in confirming the diagnosis (17 days). Forty-four patients (73%) developed ulcers typical of cutaneous leishmaniasis. Sixteen additional patients (27%) had atypical macular, papular, squamous, verrucous or acneiform skin lesions that were diagnosed only because leishmanial cultures were obtained. Of the 59 patients treated with pentavalent antimonial drugs, only 34 (58%) were cured after the first course of treatment. Lesions which were at least 2 cm in diameter, ulcerated, or caused by Leishmania braziliensis were less likely to be cured after a single course of treatment than were lesions smaller than 2 cm, nonulcerated or caused by Leishmania mexicana or Leishmania donovani.

Mouse foot-pad inoculation as an aid to the isolation of Leishmania spp. from patients

Portions of splenic or subcutaneous saline aspirates from suspected visceral or cutaneous leishmaniasis patients were inoculated into NNN media with an overlay of Schneiders medium or Schneiders medium alone for routine parasitological diagnosis. The remaining portions of the aspirates were used for preparing Giemsa-stained smears and for subcutaneous inoculation into hind foot-pads of Balb/c mice. Saline aspirates obtained from the foot-pads 2-14 d after inoculation were inoculated into Schneiders medium and examined for promastigotes. Parasite isolation was achieved from 90% of confirmed leishmaniasis patients by either culture method alone. Mouse foot-pad aspiration demonstrated parasites in 95% of all patients, and in over 80% of the confirmed cases of leishmaniasis. Combined culturing and aspirate smear examination was more efficient than foot-pad inoculation alone for the demonstration of leishmanial infection. Foot-pad aspiration does not entail killing animals and was sensitive for parasite isolation; it may be a useful short-term adjunct to existing parasite isolation methods, especially under field conditions where the risks of culture contamination may be high.

American Cutaneous Leishmaniasis

ABSTRACT: A lesion of New World leishmaniasis. evident on the palm of the hand, a rare and unusual location, is described. Routine investigations for bacteria and mycobacteria were misleading. Specific culture for Leishmania led to the correct diagnosis. Successful treatment was achieved with a pentavalent antimonial (sodium stibogluconate). The authors suggest various patient management precautions for patients with active lesions.