HIV and the transmission of Leishmania - WHO...NEWLY ACQUIRED VERSUS probabilities thatclinicalVLinaHIV-positive RE-ACTIVATED VISCERAL individual is (1) the result of a newly acquired

HIV and the transmission of Leishmania

R MOLINA L GRADONIdagger and J ALVAR

WHO Collaborating Centre for Leishmaniasis Servicio de Parasitologotilde a Centro Nacionalde Microbiologotilde a Instituto de Salud Carlos III Carretera MajadahondandashPozuelo Km 228220 Majadahonda Madrid SpaindaggerLaboratorio di Parassitologia Istituto Superiore di Sanita Viale Regina Elena 299 00161Rome Italy

Received and accepted 30 May 2003

In many countries LeishmaniaHIV co-infection is now changing the epidemiology of visceral leishmaniasis Thelevels of transmission of the parasites causing such leishmaniasis were previously dependent on the conventionalzoonotic cycle in which sand ies transmitted the parasites from infected canids to other canids or humans Theco-infection however has led not only to marked increases in the sand y transmission of the parasites from immuno-depressed individuals directly to other humans but also probably to arti cial transmission between immunodepressedintravenous-drug users as the result of needle sharing

The epidemiologyof human leishmaniasesand The zoonotic form of VL is caused bythe biology of the phlebotomine sand ies that Leishmania infantum (=Leishmania chagasi)transmit the causative parasites are naturally with infection predominantly in wild andclosely linked (Killick-Kendrick 1990) There domestic canids and occasionally in humansis little evidence to indicate that immuno- This form which is mainly endemic extendscompetent humans can develop leishmaniasis from the Mediterranean through Central Asiaexcept as a consequence of being bitten into China and also exists in Latin Americaby infected sand ies although the parasites Several species of the genera Phlebotomusmay occasionally be transmitted as the result (Old World) and Lutzomyia (New World)of blood transfusions (Cohen et al 1991 have been incriminated in the transmissionGrogl et al 1993 Singh et al 1996 Kubar of Le infantumet al 1997 Luz et al 1997 Le Fichoux The anthropondashzoonotic form caused byet al 1999 Otero et al 2000) The results Le donovani or Le infantum occurs in Eastof recent xenodiagnostic tests with sand ies Africa and the southndashwest of the Arabianindicate that the amastigotes in samples of peninsula It is normally endemic but canblood (taken from cases of LeishmaniaHIV cause extensive epidemics in humans Trans-co-infection) remain viable for at least 8 days mission of the parasites that cause this formafter the blood has been collected (R Molina of VL is predominantly human to human viaunpubl obs) non-synanthropic vectors such as P martini

From an epidemiological standpoint and P orientalis although there may alsothree main forms of human visceral leish- be other mammals acting as lsquoreservoirrsquomaniasis (VL) have been identi ed zoonotic hostsanthropondashzoonotic and anthroponotic The anthroponotic form caused by Le

donovani occurs on the Indian sub-continentIt is normally endemic but severe epidemicsReprint requests to R Molina

E-mail rmolinaisciiies fax + 34 91 5097034 can develop among humans The parasite is

DOI 101179000349803225002516

Annals of Tropical Medicine amp Parasitology Vol 97 Supplement No 1 S29ndashS45 (2003)

copy 2003 The Liverpool School of Tropical Medicine

30 MOLINA ET AL

transmitted from human to human by the and Spain give positive results in leishmaninskin tests (Pampiglione et al 1975 1976bite of a species of sand y that is strictly

peridomestic P argentipes Gramiccia et al 1990 Meller-Melloul et al1991 Marty et al 1992 Arbaji et al 1993Since the global epidemic of HIV infection

began there has been a steady rise in the Acedo-Sanchez et al 1996 Alvar et al1996 Morillas et al 1996) many detectedannual number of reported new cases of VL

associated with the virus LeishmaniaHIV cases of LeishmaniaHIV co-infection in thesecountries are probably the result of latentco-infection is emerging as a new and worry-

ing disease particularly in southndashwestern Leishmania infections that have been activatedor re-activated as the later HIV infectionEurope (Desjeux 1998) In Spain mdash the

country reporting the highest incidence of causes immunosuppression Many now thinkthat Leishmania spp should be consideredthe co-infection mdash 68 of the cases are

intravenous-drug users (Desjeux and Alvar as opportunistic pathogens in HIV-infectedpatients (Alvar et al 1992) Even in the2003) In France Italy Portugal and Spain

the outbreak of HIV infection has changed absence of HIV Le infantum infectionsmay be re-activated following immunologicalthe epidemiology of VL from a disease pre-

dominantly found in children to one more distress (Ma et al 1979 Kubar et al 1998)If the natural history of VL in HIV-common in adults (WHO 1999) HIV co-

infection may re-activate latent leishmanial positive individuals is to be fully understoodlong-term longitudinal immunological andinfections increase the level of transmission

of Leishmania (particularly humanndashhuman parasitological investigations on cohorts ofindividuals at risk of both HIV and Leishmaniatransmission) by phlebotomine sand ies and

faciliate arti cial transmission of Leishmania infection will have to be conducted As theincidence of leishmaniasis in the areas mostvia the sharing of contaminated syringes and

needles from one intravenous-drug user aVected by the co-infection is relatively lowthe cohorts studied will have to be large The(IVDU) to another (see below)relevant epidemiological data already availablemay be suYcient however to estimate the

NEWLY ACQUIRED VERSUS probabilities that clinical VL in a HIV-positiveRE-ACTIVATED VISCERAL individual is (1) the result of a newly acquired

LEISHMANIASIS THE ITALIAN leishmanial infection or (2) the result ofEXAMPLE a re-activated latent infection In 1998

L Gradoni and A Scalone attempted toestimate these two probabilities in SicilyIt is estimated that only one in every ve to

10 immunocompetent individuals who have by assimilating and updating the relevantepidemiological-surveillance data and thenbeen infected with Le donovani or Le infantum

ever develops clinical VL (Desjeux 1992 using the resultant data-set in an epidemio-logical model (unpubl obs) Their analysisBadaro et al 1996) Since a T-lymphocyte-

mediated immune response is necessary to of the natural history of the LeishmaniaHIVco-infection is described below in extensocontrol Leishmania infection however those

residents of endemic areas who have beenmade immunode cient as the result of HIVinfection are far more prone to VL than their Surveillance Methodology

In 1989 a programme based on the activeimmunocompetent neighbours Strains ofLeishmania that are usually non-pathogenic detection of LeishmaniaHIV co-infection was

implemented in collaboration with some 148in humans and even lower trypanosomatids(Chicharro and Alvar 2003) may all cause infectious-disease units sited in universities

and hospitals throughout Italy (Gradoni et alillness in HIV-positive individuals As manyasymptomatic individuals in Italy France 1996) The units selected from the 260

HIV AND Leishmania TRANSMISSION 31

clinical diagnostic centres which were then The Situation in ItalyBetween 1985 and 1997 183 cases ofrecording AIDS cases were chosen because

they were not only reporting relatively high LeishmaniaHIV co-infection were recordedin Italy Most (68) of the co-infectionsnumbers of HIV-positive patients but were

also in regions where VL was endemic were in patients who ful lled the criteria forAIDS In areas of Italy where leishmaniasisCollaborators were asked to provide serum

bone-marrow peripheral-blood or skin-biopsy was endemic the incidence of VL amongHIV-positive residents (16 cases100) wassamples from any patient who when sero-

positive for HIV or suVering from AIDS about 500 times higher than that among theirHIV-negative neighbours In four distinctdeveloped an infection that was clinically

suspected to be VL One or more of the lsquohotspotsrsquo 12ndash100 of the HIV-positivesdeveloped VL The annual number of co-following clinical and laboratory ndings were

considered indicative of VL fever of unknown infection cases showed a sharp increase in1991 but then stabilized at about 21 casesorigin splenomegaly hepatomegaly hyper-

gammaglobulinaemia and pancytopenia year (Fig 1)The presence of single or multiple nodularulcerative skin lesions was considered indi-cative of cutaneous leishmaniasis (CL) A Epidemiological Model

A model was constructed to test two alternativeretrospective survey was conducted to identifyany cases of co-infection that had presented hypotheses that the VL seen in HIV-positives

represented (1) newly acquired infectionsat the collaborating units before 1989 con- rmatory diagnosis was then based on the or (2) re-activation of latent leishmanial

infectionslaboratory examination of stored sera andbone-marrow smears Possible Leishmania To test the rst of these hypotheses the

number of expected co-infections was derivedinfections diagnosed in AIDS patients wereroutinely reported through the national by applying a known force of leishmanial

infection ( p see below) to an estimated HIV-system of AIDS-case noti cation with otherassociated pathologies (ie those not indi- infected population exposed to Leishmania

during each Leishmania-transmission seasoncative of leishmaniasis) to the National AIDSRegistry It was assumed that the level and mode

FIG 1 Temporal distribution of the cases of Leishmania co-infection among those with AIDS (f ) and otherHIV-positive individuals (e ) recorded during epidemiological surveillance in Italy

32 MOLINA ET AL

of Leishmania transmission to and among the number of HIV-positive residents of Sicilywho lived in areas where leishmaniasis wasHIV-positive individuals were the same asendemicthose to and among the HIV-negatives that

each HIV-positive individual infected withLeishmania developed clinical VL and that

force of infection ( p)p had been constant during the 10-year studyThe force of infection (Lysenko and Beljaevperiod (1985ndash1994)1987) was estimated as the mean numberTo test the second hypothesis the numberof times a resident of Sicily was inoculatedof expected co-infections was calculated bywith Leishmania over a year from LSTapplying an estimate of the annual incidencedata indicating the prevalence of leishmanialof HIV infection to a population that includedinfection among Sicilans aged 0ndashl5 years whomany individuals mdash the number being indi-were assumed to be non-immune Surveyscated by the results of leishmanin skin testsbased on LST have been carried out in Sicily(LST) mdash who were asymptomatic carrierssince 1975 Their results indicate that theof Leishmania It was assumed that eachforce of infection in Sicily varies little fromindividual found positive in an LST had ayear to year and is similar to those in otherlatent leishmanial infection that would developleishmaniasis-endemic areas of southern Italyinto clinical VL following that individualrsquos(Pampiglione et al 1975 Gradoni et alinfection with HIV and that the age-related1993 unpubl obs)prevalence of LST positivity was constant

over the study period

the population potentially harbouring

LeishmaniaSources of Data

The results of the LST surveys in Sicilywere also used to estimate the number ofprevalence and incidence of HIVindividuals on the island who were LST-infectionpositive and therefore possibly carrying latentThe prevalence and incidence of HIVLeishmania infections Particular attentioninfection in Italy in 1994 were estimatedwas paid to the LST results for residents ofusing mathematical models lsquoback-calculationrsquoSicily who were aged 20ndash30 years this age-the annual numbers of AIDS cases noti edgroup being considered the one at greatestand the results of pilot studies on HIV sero-risk of HIV infectionconversion (Pezzotti et al 1995 Rezza 1998)

Prevalence was estimated by subtracting thenumber of HIV-attributable deaths prior to

reported incidence of the1994 from the accumulative incidence of HIVLeishmaniaHIV co-infectioninfection Annual incidence represented allThe annual numbers of cases LeishmaniaHIVthe new HIV infections that occurred inco-infection detected among Sicilian residents1994during epidemiological surveillance betweenGradoni et al (1996) used maps of the1985 and 1994 were compared with thosedistribution of leishmaniasis in Sicily (basedpredicted using each of the two mathematicalon accurate locality data for all cases ofmodelshuman and canine leishmaniasis that had

been recorded over the previous 18 years)to estimate the size of the human population

Evaluation of the Modelsat risk of Leishmania infection on the islandThey then used the same maps and the hypothesis 1 (lsquonew infectionsrsquo)relevant AIDS-case noti cation reports stored In Sicily the number of HIV-positives living

in areas where VL was endemic ranged fromat the National AIDS Registry to estimate


approximately 800 in 1985 to approximately in the period 1986ndash1989 to 16ndash24 casesyearwith a total of 376 cases over the study4200 in 1994 (Fig 2) The corresponding

numbers of AIDS cases showed the same periodtrend (Fig 2) The mean force of leishmanialinfection in these areas was calculated to be

observed HIVVL cases0003 infectionperson-year indicating thatThe actual recorded numbers of HIV-the areas are only hypo-endemic for VL Bypositive individuals found to have VL inapplying this p-value to the number of HIV-Sicily increased from nil in 1985 to nine inpositives present in the areas of the island1994 with a total of 35 HIVVL cases overwhere VL is endemic the annual numbersthe study period The trend in the numbersof cases of co-infection were predicted toof reported cases was similar to that pre-increase from two in 1985 to 12 in 1994dicted when the lsquonew-infectionrsquo hypothesiswith a total of 74 cases over the study periodwas applied and diVered signi cantly fromthat predicted when the lsquore-activationrsquo hypo-

hypothesis 2 (lsquore-activationsrsquo) thesis was assumed to be correct (Plt001The estimated incidence of HIV infection in Fig 3)Sicily peaked at about 600ndash700 casesyearbetween 1986 and 1989 and then decreasedto about 200ndash300 casesyear (Fig 2) Most Discussion

In Italy the problem of HIVVL co-infectionof these cases were aged 20ndash30 years A meanof 80 of the Sicilian residents aged 20ndash30 is no longer as alarming at it appeared to be

in the early 1990s when the number of casesyears who were checked in LST surveys werefound LST-positive By applying this pro- was showing dramatic increases from one

year to the next Although in some areas ofportion to the size of the HIV-positive popu-lation the numbers of cases of co-infection the country the incidence of VL does appear

to have increased recently this upward trendwere predicted to decrease from 48ndash56year

FIG 2 AIDS incidence (f ) and the estimated trends of HIV prevalence (e ) and incidence (^) in leishmaniasis-endemic areas of Sicily

34 MOLINA ET AL

FIG 3 The numbers of cases of visceral leishmaniasis observed among the HIV-positive residents of Sicilybetween 1985 and 1994 (f ) and the numbers expected if hypothesis 1 (^ lsquonewly acquired leishmanial infectionsrsquo)or hypothesis 2 (e lsquore-activations of latent leishmanial infectionsrsquo) were correct

appears to be unrelated to HIV (Gradoni who have frequently taken their vacations inregions of the Mediterranean littoral whereet al 1996) It seems that the predicted

ood of HIVVL cases has been stemmed the disease is endemicThe epidemiological models have to beby the largely successful introduction of anti-

retroviral treatment for those found sero- treated with some caution as they may be toosimplistic Although the two hypotheses werepositive for HIV The quarterly incidence of

AIDS in Italy peaked at 10 cases100000 assumed to be mutually exclusive both newinfections and re-activations may contributeinhabitants in 1995 falling to six cases

100000 in the rst quarter of 1998 to the actual number of clinical HIVVL casesAlthough the predicted number of cases forThe results of the epidemiological modelling

discussed above indicate that in Sicily at each year was compared with the actualnumber of cases diagnosed in the same yearleast most cases of HIVVL co-infection are

the result of individuals who are already HIV- an incubation period mdash the length of whichis diYcult to estimate mdash will have separatedpositive being newly infected with Leishmania

Had such co-infection been predominantly co-infection from diagnosis Some of theassumptions and estimates made appear morethe result of those with (latent) leishmanial

infection being infected with HIV then many reasonable than others The estimates of pand of the proportion of the population whomore cases should have been detected in the

rst few years of the present study period may have latent Leishmania infections forexample were both based on LST dataas HIV swept through Sicily Such large

numbers of cases were not observed at that showed consistency over a long periodThese estimates were supported by a stablethis time in Sicily in other areas of the

Mediterranean basin where VL is endemic incidence of recorded VL among the immuno-competent population of Sicily during theor in the young people of northern Europe


study period in the absence of measures to increases the risk of VL by 100- to 1000-fold in areas where VL is endemic (WHOcontrol the canine reservoir or phlebotomine

vectors and of dramatic changes in local 1999) In western Europe IVDU who sharesyringes form the main population at risk ofenvironmental conditions (Cascio et al

1997) The possibility that HIV infection HIV infection and account for 44 of theHIVAIDS cases (Desjeux 1998) In a jointhas led to or is associated with unusual

routes of leishmanial infection (see below) consultative meeting on the LeishmaniaHIVco-infection held in September 1998 it waswas not considered in the epidemiological

models established that IVDU had a 25- to 36-foldhigher risk of VLAIDS than other AIDScases (WHO 1998) In Spain and Italy

ALTERNATIVE ANTHROPONOTIC biochemical variants of Le infantum thatCYCLES FOR Leishmania infantum rarely if ever cause leishmaniasis in immuno-

competent patients have been recoveredfrom cases of the HIVLeishmania co-infectionIn the predominant natural cycle of trans-(Gramiccia et al 1995 Jimenez et al 1995mission an individual whether HIV-positivePratlong et al 1995 Agostoni et al 1998)or HIV-negative becomes infected with LeAmastigote-infected macrophages can beinfantum when he or she is bitten (usuallyfound in approximately 50 of bloodsmearsin rural or peri-urban settings) by a sand yfrom co-infected patients (Mart otilde nez et althat has already been infected when taking1993 Medrano et al 1993) and pro-a bloodmeal from an infected canid Themastigotes can be found in 67 of NNNpossibility of other routes of transmissioncultures of buVy coats from such patientssuch as needle-mediated infection should(Lopez-Velez et al 1995)not however be ruled out (Alvar et al

The AIDS-case surveillance system that1992) Direct humanndashsand yndashhuman trans-covers the Madrid region of Spain has alsomission may also occur particularly fromprovided data indicating that needle-sharingHIV-positives who carry particularly highcarries a risk of HIVLeishmania co-infectionnumbers of leishmanial amastigotes in their(Amela et al 1996) As this system onlyperipheral bloodrecords AIDS cases VL diagnoses in anyHIV-infected individuals who fail to meetthe diagnostic criteria for AIDS are notAn Arti cial Cycle of Transmission

It seems likely that not only HIV but also included In total 6652 cases of AIDS werediagnosed between 1982 and 1993 andLeishmania may be transmitted on and in

shared needles and syringes among IVDU recorded by the Madrid system Only 166(25) of these cases developed VL andThe observation that mammals may be

infected with Leishmania either accidentally the dates of diagnosis of both the AIDSand VL were available for only 137 of the(Owens et al 2001) or experimentally

(Palatnik de Sousa et al 1996) by trans- co-infection cases The VL was diagnosedbefore the AIDS in 33 cases after the AIDSfusions of blood from infected hosts under-

lines the possibility of this mode of trans- in 65 cases and at the same time as theAIDS in 39 cases The prevalence of VLmission There has also been at least one

case of laboratory-acquired Le donovani among the AIDS cases who were IVDU wassigni cantly higher than that in any of theinfection as the result of a needlestick injury

(Freedman et al 1987) Although there is other exposure-groups (relative risk=25795 con dence interval=164ndash401)as yet no direct evidence of the spread of

Leishmania through the sharing of syringes In Italy the unexpected occurrence ofsmall hotspots of HIVLeishmania co-infectionamong IVDU there is much indirect evi-

dence indicating that this happens AIDS in which every detected case of HIV infection

36 MOLINA ET AL

may have VL also indicates that transmission Europe between 1990 and 1998 the pre-dominance of IVDU who were identi edis not restricted to the normal canidndash

sand yndashhuman route The observations that as the main population at risk was clear(WHO 1999)IVDU represent 84 of the Italian cases of

HIVLeishmania co-infection but only 64 ofItalian AIDS patients without leishmaniasis

Natural Humanndashsand yndashhuman(64) and that the zymodeme spectrum of

Transmissionthe Le infantum from the co-infected cases

Most humans are infected with Le infantumdiVers from that of the parasitic isolates

when they are bitten by sand ies that havefrom HIV-negative adults and the distances

already been infected when they fed on dogsbetween the hotspots also support the view

or other canids harbouring the parasitethat leishmanial parasites can be transmitted

As uninfected sand ies are not frequentlyon needles and syringes (Gradoni et al

infected with Le infantum as they feed on1996) In northern Italy 17 (77) of the 22

immunocompent humans carrying the para-co-infected patients investigated by Agostoni

site humanndashhuman transmission of Leet al (1998) were IVDU

infantum via a sand y is rare There isPineda et al (1998) investigated the

however considerable evidence indicatingfactors associated with VL in individuals

that uninfected sand ies feeding on indi-infected with HIV-1 who lived in southern

viduals co-infected with Le infantum andSpain When Giemsa-stained smears of bone-

HIV are quite likely to become infected withmarrow aspirates from 291 HIV-1 carriers

the parasite Transmission of leishmanialwere examined 45 were found positive for

parasites from a HIV-positive human is there-amastigotes Thirty-two of the amastigote-

fore more likely than transmission from apositive carriers of HIV-1 had symptomatic

HIV-negative individual Most of the dataVL the other 13 having subclinical infections

indicating that sand y-mediated transmissionwith Leishmania Symptomatic VL again

of Le infantum from co-infected HIV-positiveappeared particularly common among the

individuals occurs have been collected duringHIV-positive IVDU included in the study

xenodiagnostic tests with sand iesAlthough this association was not quite foundto be statistically signi cant when adjust-ments were made for clinical category and indirect xenodiagnosis

The con rmation of a suspected diagnosis ofgender in a multivariate analysis Pinedaet al (1998) thought that Leishmania trans- leishmaniasis in immunodepressed patients

frequently requires the use of several tech-mission through the sharing of needles wasvery probable The isolations of the MON-18 niques Although the examination of bone-

marrow aspirates for amastigotes has beenzymodeme of Le donovani from a Portuguesedrug addict with clinical VL and AIDS proposed as the best technique it requires

an invasive and painful procedure and is not(Campino et al 1994) and of the MON-253 zymodeme of Le infantum from a cluster highly sensitive In the search for a dia-

gnostic test that is less invasive and less pain-of three co-infected patients all IVDUliving in the same town in northndasheastern ful for patients who may already be seriously

ill the use of indirect xenodiagnosis (IXD) mdashSpain (Chicharro et al 1999) also supportthe view that leishmanial amastigotes are that is the feeding of uninfected (usually

laboratory-bred) sand ies through a mem-being transmitted on shared needles WhenCruz et al (2002) checked syringes discarded brane on a sample of venous blood from

the suspected case of leishmaniasis mdash hasby IVDU in Madrid they found 34ndash52to be PCR-positive for leishmanial DNA been considered Molina et al (1992) found

IXD with Phlebotomus perniciosus an importantIn a retrospective analysis of 965 casesof co-infection reported in southndashwestern vector of Le infantum in the southndashwestern


Mediterranean region very useful in con- Pakistan have led to the hypothesis that skin rming that an AIDS patient had VL parasites rather than parasites circulating inSubsequently Molina et al (1994) investi- blood monocytes might be the principalgated the potential usefulness of IXD in the source of infection for sand ies in areasroutine detection of leishmaniasis in those where anthroponotic leishmaniasis is foundco-infected with Le infantum and HIV (Rab et al 1992)]and attempted to establish a standard pro- Using IXD parasites can be clearlytocol The study designed to investigate the observed 48 h after sand y feeding parasitesinfectivity to P perniciosus of 10 HIV-positive in cultures of aspirates and blood usuallyindividuals who had symptoms indicative of take longer to multiply suYciently for themVL (N=9) or had duodenal amastigotes to be detected Although IXD is very usefuldetected during routine endoscopy ( N=1) for detecting Le infantum in HIV-positivealso served to compare the IXD with the patients its application is for the momentmore conventional methods of diagnosis largely restricted to those cases who haveThe other methods tested in parallel with given negative results in other diagnosticthe IXD were IFAT for the detection of tests even though there is strong clinical evi-anti-Leishmania antibodies the microscopical dence indicating that they have leishmaniasisexamination of Giemsa-stained smears of IXD is a highly sensitive technique whenbone-marrow aspirates and peripheral blood used on immunodepressed patients Molinaand NNN cultures of bone-marrow aspirates et al (1998) for example used IXD toand peripheral mononuclear cells Records demonstrate the presence of viable parasiteswere made for each patient of associated in 21 (95) of 22 blood samples from casesinfections fever (for gt2 weeks) spleno- of Le infantumHIV co-infection and in bloodmegaly treatment for the leishmaniasis and

samples from two other patients who thoughthe response observed the sand y-feeding

HIV-negative were immunodepressed as theprocedure and other information (Table 1)

result of acute lymphoblastic leukaemiaThe P perniciosus used came from a local

The observation that blood from caseslaboratory colony kept at 27plusmn1degC andof LeishmaniaHIV co-infection remains90ndash100 relative humidity with a 17-h-infective to the sand ies used in IXD evenlight7-h-dark photoperiod (Molina 1991)if used after storage for at least 8 daysBatches of 30ndash150 female P perniciosus(unpubl obs) adds support to the idea thatcollected 4ndash21 days post-eclosion were fedleishmanial infections can be transmitted byin the presence of some males through ablood transfusionmembrane on anticoagulated peripheral blood

IXD has uses in the eld of epidemiologyfrom each patient The bloodfed sand iesother than the demonstration of leishmanialwere dissected 2ndash7 days after taking theirco-infection in those who are HIV-positivebloodmeals so that their guts could be checkedIt has been used for example to show thefor promastigotes All 10 of the patients wereimportance of haematogenic disseminationfound positive by the IXD but only eightin the development of mucosal leishmaniasisseven ve and ve respectively had positiveand other metastatic manifestations of Lecultures of bone-marrow aspirates positivebraziliensis infection (Mart otilde nez et al 1992)cultures of peripheral blood monocytesDa-Cruz et al (1992) used IXD to study theamastigotes detected in their bloodsmearsfailure of T-cell-mediated immune responsesor were found seropositive in the IFATto prevent diVuse cutaneous leishmaniasis(Table 1) Even in the absence of cutaneousdeveloping in HIV-positives infected withmacrophages carrying amastigotes mostLe braziliensis As the result of using(up to 93) of the sand ies taking a blood-P argentipes in IXD Addy and Nandymeal were infected [The results of studies

conducted on Le infantum-related VL in (1992) concluded that the skin parasites in

38 MOLINA ET AL

TA

BL

E1

C

linic

alan

dpa

rasi

tolo

gica

lda

tafo

r10

case

sof

Lei

shm

ania

infa

ntu

mH

IVco

-inf

ecti

onin

vest

igat

edby

indi

rect

xeno

diag

nosi

s

Cas

e

12

34

56

78

91

0

Sex

MM

MF

MM

MM

MM

Age

(yea

rs)

36

333

525

382

922

2828

37

Exp

osu

regr

oup

for

HIV

infe

ctio

nIV

DU

IVD

UIV

DU

IVD

UH

SIV

DU

IVD

UIV

DU

IVD

UIV

DU

Ass

ocia

ted

infe

ctio

n(s

)P

TB

O

CC

MV

PC

PE

CP

TB

E

CO

CE

PT

BN

oN

oP

CP

Fev

er

Yes

No

Yes

Yes

Yes

Yes

Yes

Yes

Yes

Yes

Spl

enom

egal

yY

esN

oY

esY

esY

esY

esY

esY

esY

esN

oL

euco

cyte

cou

nt

(leu

cocy

tes

ml)

270

02

400

NA

250

043

0036

0024

00

1600

190

059

00C

D4

+co

un

t(C

D4

+ce

llsm

l)4

010

68

5648

51

144

201

48N

DH

aem

oglo

bin

(gl

)8

48

3N

A7

28

49

10

110

65

74

Pla

tele

tco

un

t(p

late

lets

nl)

71

164

NA

8475

98

7317

91

311

70A

mas

tigo

tes

inb

lood

smea

rN

oY

esN

oY

esY

esY

esN

oN

oN

oY

esA

mas

tigo

tes

insm

ear

ofbo

ne-

mar

row

aspi

rate

N

oY

esY

esY

esY

esY

esN

oN

oN

oY

esS

erol

ogy

(IF

AT

titr

e)1

160

lt1

40

18

0lt

140

lt1

40lt

140

116

01

320

140

lt1

40B

one-

mar

row

cult

ure

++

++

++

shy+

shy+

Per

iphe

ral-

bloo

d-m

onoc

yte

cult

ure

shy+

++

shy+

++

shy+

Tre

atm

ent

for

leis

hm

ania

sis

(day

s)2

00

00

0R

elap

seR

elap

seR

elap

se0

Res

ult

ofin

dire

ctxe

nod

iagn

osis

++

++

++

++

++

Blo

odst

orag

eat

4degC

pri

orto

xen

odia

gnos

is(h

)6

62

46

366

2448

722

4

ofb

lood

fed

san

d

ies

fou

ndin

fect

ed22

837

514

262

79

29

48

38

010

713

853

3

M

Mal

eF

fe

mal

eIV

DU

in

trav

enou

s-dr

ug

use

rH

S

hete

rose

xual

P

TB

p

ulm

onar

ytu

berc

ulos

is

OC

or

alca

nd

idia

sis

CM

V

cyto

meg

alov

iru

sin

fect

ion

P

CP

P

neum

ocys

tisca

rini

iE

PT

B

extr

apu

lmon

ary

tub

ercu

losi

sN

A

not

avai

lab

le

ND

n

otd

one


a patient with nodulo-ulcerative post-kala-azar HIV-positive individual who is co-infectedwith Le infantum raises the possibility thatdermal leishmaniasis represented a source of

infection in a VL focus in West Bengal the cases of co-infection (and any otherimmunocompromised cases of VL) may actAfter several years of using IXD to

test blood samples from Spanish cases of as secondary reservoirs of VL in a natural butanthroponotic cycle of the disease DirectLe infantumHIV co-infection Molina et al

(1996) developed a standard protocol xenodiagnosis (DXD) mdash in which uninfectedsand ies are allowed to feed directly onSamples of peripheral blood are collected in

tubes with anticoagulant (usually heparin humans mdash has recently been used to explorethis possibility (Molina et al 1999) Eachalthough EDTA or sodium citrate are equally

satisfactory) and brought to the laboratory of six co-infected patients was asked to placeone of his or her hands into a small cagerefrigerated at 4degC Each is tested by oVer-

ing 50 7-day-old laboratory-bred female containing 25 7-day-old laboratory-bredfemale P perniciosus (and a similar numberP perniciosus a 15-ml sample of blood held

at 37degC for 1 h in a sterile membrane- of male ies) for 15 min (Fig 4 Table 2)The unfed ies were carefully removed withfeeding apparatus using the skin of a 3-day-

old chicken as the membrane Any unfed an electrical aspirator and killed while theblood-fed females were kept in the cage for ies are then removed with an electrical

aspirator and killed with chloroform or CO2 at least 72 h before being dissected (Molinaet al 1996) All six patients were found toThe fed ies are maintained on 30 fructose

solution replaced daily for 48ndash72 h before be infective to the sand ies their infectivitybeing negatively correlated with their CD4+the dissections begin Each y to be dis-

sected is anaesthetized with CO2 placed in cell counts (Fig 5) Counts of CD4+cells

may therefore be a useful indicator of thea drop of sterile phosphate-buVered saline(PBS) on a sterile microscope slide and infectivity of a co-infected patient

To summarize the results of xeno-decapitated with sterile needles The midgutis then drawn out transferred to another diagnoses have indicated that (1) vector

sand ies are readily infected by feeding ondrop of sterile PBS covered with a sterilecoverslip and examined under the micro- immunodepressedcases of Le infantumHIV

co-infection whereas immunocompetentscope for promastigotes If a gut is found to beheavily infected it is gently ruptured by press- individuals infected with Le infantum are

poorly infective to such insects (2) CD4+ing on the coverslip and used to inoculateNNN medium so that the Leishmania strain cell counts are useful indicators of the

infectivity of co-infected patients and (3) atcan be isolated and typed by iso-enzymeanalysis If a gut is found to contain only a least some of the Le infantum in the blood

of those co-infected with HIV remain infectivefew promastigotes 48 h after the infectivefeed the surviving ies that had fed on the to sand ies for at least 8 days after the blood

has been collectedsame sample are maintained until at leastday 7 post-bloodmeal before they too aredissected in the hope that by then some willbe heavily infected and of use for parasite Epidemiological Implicationsisolation The ease with which sand ies may be

infected when fed directly or indirectly onthe blood of those with Le infantumHIVco-infection has considerable epidemiologicaldirect xenodiagnosis

The ease with which sand ies can be infected implications particularly as the cases of co-infection may remain asymptomatic the VLwith leishmanial parasites by feeding them

through a membrane on blood from a in HIV-positive individuals often responds

40 MOLINA ET AL

FIG 4 Direct xenodiagnosis of a suspected case of Leishmania infantumHIV co-infection

TABLE 2 Clinical and parasitological data for six cases of Leishmania infantumHIV co-infection investigated by directxenodiagnosis

Case

1 2 3 4 5 6

Age (years) 26 27 29 38 27 32Associated infection(s) No PCP OC T MAI OC NoFever Yes Yes Yes No Yes YesSplenomegaly Yes Yes Yes Yes No NoLeucocyte count (leucocytesml) 2200 3910 2400 4450 980 720CD4+count (CD4+cellsml) 120 4 45 12 39 28Haemoglobin (gl ) 86 87 101 96 79 87Platelet count (plateletsnl ) 125 300 72 56 125 70Serology (IFAT titre) 1640 lt140 ND ND 1320 lt140Bone-marrow culture ND ND + + + shyPeripheral-blood-monocyte culture + + ND ND ND shyResult of direct xenodiagnosis + + + + + + of bloodfed sand ies found infected 91 857 375 889 385 182

Relapse after treatment for leishmaniasisPCP Pneumocystis carinii OC oral candidiasis T toxoplasmosis MAI Mycobacterium avium infection ND not done

poorly to treatment and VL may develop at 1992 Arbaji et al 1993 Acedo-Sanchezet al 1996 Morillas et al 1996) Le Fichouxany stage of the HIV infection (Pineda et al

1998) The results of leishmanin skin tests et al (1999) detected leishmanial DNA innine blood samples (out of 76 found sero-and lymphoproliferative tests in vitro indicate

that in areas where VL is endemic many positive for Leishmania) from the Monacoblood bank indicating that Le infantumindividuals though asymptomatic may be

carrying leishmanial infections (Pampiglione circulates albeit perhaps only intermittentlyand at a low density in the blood of asymp-et al 1975 1976 Gramiccia et al 1990

Meller-Melloul et al 1991 Marty et al tomatic blood donors


FIG 5 The relationship between the infectivity of untreated cases of Leishmania infantumHIV co-infection tolaboratory-bred Phlebotomus perniciosus (e ) and the CD4+cell counts of the cases (f )

It seems that some iso-enzymatic pheno- the strictly anthroponotic VL caused by Ledonovani seen in India where even HIV-types of Le infantum which have been

detected in HIV-positive individuals but negative cases of VL are highly infective tosand ies (Theodor 1964) As most (915)not in immunocompetent humans or dogs

are particularly associated with immuno- cases of Le infantumHIV co-infectiondetected in southndashwestern Europe have lowdepression (Gramiccia et al 1995 Jimenez

et al 1995 Gradoni et al 1996 Pratlong CD4+counts of lt200 cellsml (WHO1999) many are probably infective to theiret al 1995 Rosenthal et al 1995 Harrat et al

1996 Agostoni et al 1998 Chicharro et al local sand ies The overall prevalence ofhuman co-infection in the VL-endemic areas1999) Such phenotypes occur among the

many zymodemes to be found in wild sand y of Europe is very low when compared withthat of canine leishmaniasis There maypopulations (Rioux et al 1986 Gradoni et al

1991 Martotilde n-Sanchez et al 1994 1995 be foci however in which humanndashhumantransmission via sand ies is playing an1996) and at least some of them can infect

laboratory-bred P perniciosus ( J M Lohse important part in the local epidemiology ofVL Cases of LeishmaniaHIV co-infectionand R Molina unpubl obs)

In the presence of HIV Le infantum may be clustered together for example ininstitutions specializing in the care of AIDSadapted to the peridomestic environment

could evolve without the intervention of any patients or the detoxi cation of IVDUIn some countries including Spain suchother vertebrate host apart from humans to

a true anthroponosis (Tesh 1995) In some institutions are usually in peri-urban or iso-lated rural settings in which sand ies areaspects this situation would emulate that of

42 MOLINA ET AL

often common It may only take one case of bednets and indoor insecticide spraying inhigh-risk settings (such as detoxi cation andLeishmaniaHIV co-infection to trigger anAIDS-treatment centres)outbreak of VL the 1980 outbreak of VL in

West Bengal possibly developed from onecase of post-kala-azar dermal leishmaniasis

REFERENCES(Addy and Nandy 1992)Clearly the unequivocal demonstration

of immunode cient-humanndashsand yndashhuman Acedo Sanchez C Martotilde n Sanchez J VelezBernal I D Sanchotilde s Marotilde n M C Louassini Mtransmission will be diYcult in an environ-Maldonado J A amp Morillas Marquez F (1996)ment in which canidndashsand y transmission isLeishmaniasis eco-epidemiology in the Alpujarra

relatively common It may be possible to region (Granada province southern Spain) Inter-demonstrate a new leishmanial infection in national Journal for Parasitology 25 303ndash310

Addy M amp Nandy A (1992) Ten years of kala-azaran individual who is not an IVDU but livesin West Bengal Part I Did post-kala-azar dermalnear a case of Le infantumHIV co-infectionleishmaniasis initiate the outbreak in 24-Parganasand distant from any infected canids EvenBulletin of the World Health Organization 70 341ndash346

then it would be almost impossible to prove Adler S amp Theodor O (1931) A study of the sand ypopulation in endemic foci of infantile kala-azar inthat there are no infected canids nearbyItaly Bulletin of Entomological Research 22 105ndash113and the possibility of transmission from

Adler S amp Theodor O (1935) Investigations onan immunocompetent human carrier of LeMediterranean kala-azar VIII Further observations

infantum could not be excluded (Le Fichoux on Mediterranean sand ies Proceedings of the Royalet al 1999) Society of London Series B 116 505ndash515

Agostoni C Dorigoni N Mal tano A Caggese LThe World Health Organization estimatesMarchetti G Corona S Gatti S amp Scaglia Mthat in southndashwestern Europe 769 of the(1998) Mediterranean leishmaniasis in HIV-infectedcases of co-infection are aged 31ndash50 yearspatients epidemiological clinical and diagnostic

711 of them are IVDU aged gt15 years features of 22 cases Infection 26 93ndash98Alvar J Gutierrez-Solar B Molina R Lopez-and up to 9 of people with AIDS suVer

Velez R Garcotilde a-Camacho A Martotilde nez Pfrom VL (Desjeux 1998 WHO 1999)Laguna F Cercenado E amp Galmes A (1992)The number of co-infected individuals isPrevalence of Leishmania infection among AIDS

expected to fall in Europe as the result of patients Lancet 339 1427the widespread use of anti-HIV drugs and Alvar J Gutierrez-Solar B Pachon I Calbacho E

Ram otilde rez M Valles R Guillen J L Canavate Cto rise in South Asia and sub-Saharan Africaamp Amela C (1996) AIDS and Leishmania infantum(Desjeux 1998 WHO 1999) Discouragingnew approaches for a new epidemiological problem

the practice of syringe-sharing among IVDUClinics in Dermatology 14 541ndash546

may further reduce the incidence of the Amela C Lopez-Gay D Alberdi J C amp Castilla J(1996) Injecting drug use as risk factor for visceralLeishmaniaHIV co-infection although it willleishmaniasis in AIDS patients European Journal ofhave little if any eVect on the leishmanialEpidemiology 12 91ndash92infection of sand ies feeding on the cases of

Arbaji A K Gradoni L amp Gramiccia M (1993)co-infection The potential threat posed to Leishmanin skin test survey in a focus of highthe rest of their communities by cases of endemicity of Leishmania major in Jordan Acta

Tropica 54 77ndash79co-infection as sources of sand y infectionBadaro R Jones T C Carvalho E Mwill be diYcult to reduce Several control

Sampaio D Reed S G Barral A Teixeira R ampmeasures might be necessary such as Johnson Jr W D (1996) New perspectives on aperiodical counts of CD4+cell counts (allow- subclinical form of visceral leishmaniasis Journal of

Infectious Diseases 154 1003ndash1011ing the epidemiological risk posed by eachCampino L Santos-Gomes G Pratlong Fcase of co-infection to be evaluated) the

Dedet J P amp Abranches P (1994) The isolationrapid and prolonged antileishmanial treat-of Leishmania donovani MON-18 from an AIDS

ment and isolation of co-infected patients patients in Portugal possible needle transmissionParasite 1 391ndash392and the use of insecticide-impregnated


Cascio A Gradoni L Scarlata F Gramiccia M in HIV-1-infected individuals in Italy AIDS 10785ndash791Giordano S Russo R Scalone A Camma C

amp Titone L (1997) Epidemiologic surveillance of Gramiccia M Bettini S Gradoni L Ciarmoli PVerrilli M L Loddo S amp Ciccalo C (1990)visceral leishmaniasis in Sicily Italy American Journal

of Tropical Medicine and Hygiene 57 75ndash78 Leishmaniasis in Sardinia 5 Leishmanin reaction inthe human population of a focus of low endemicityChicharro C amp Alvar J (2003) Lower trypano-

somatids in AIDS patients Annals of Tropical of canine leishmaniasis Transactions of the RoyalSociety of Tropical Medicine and Hygiene 84 371ndash374Medicine and Parasitology 97 (Suppl 1) 000ndash000

Chicharro C Sirera G Ares M Sans A Videla S Gramiccia M Gradoni L amp Troiani M (1995)Heterogeneity among zymodemes of Leishmaniaamp Alvar J (1999) Is Leishmania infantum zymodeme

MON-253 involved in an outbreak among intra- infantum from HIV-positive patients with visceralleishmaniasis in south Italy FEMS Microbiologyvenous drug users Transactions of the Royal Society

of Tropical Medicine and Hygiene 93 385ndash386 Letters 128 33ndash38Grogl M Daugirda J L Hoover D L Magill A JCohen C Corazza F De-Mol P amp Brasseur D

(1991) Leishmaniasis acquired in Belgium Lancet amp Berman J D (1993) Survivability and infectivityof viscerotropic Leishmania tropica from Operation338 128

Cruz I Morales M A Rodr otilde guez A Noguer I amp Desert Storm participants in human blood productsmaintained under blood bank conditions AmericanAlvar J (2002) Leishmania in discarded syringes

from intravenous drug users Lancet 359 1124ndash1125 Journal of Tropical Medicine and Hygiene 49 308ndash315Harrat Z Pratlong F Belazzoug S Dereure JDa-Cruz A M Machado E S Menezes J A

Rutowitsch M S amp Coutinho S G (1992) Deniau M Rioux J A Belkaid M amp Dedet J P(1996) Leishmania infantum and Leishmania major inCellular and humoral immune responses of a patient

with American cutaneous leishmaniasis and AIDS Algeria Transactions of the Royal Society of TropicalMedicine and Hygiene 90 625ndash629Transactions of the Royal Society of Tropical Medicine

and Hygiene 86 511ndash512 Jimenez M I Ferrer-Dufol M Canavate CGutierrez-Solar B Molina R Laguna F Lopez-Desjeux P (1992) Human leishmaniases epidemiology

and public health aspects World Heath Statistics Velez R Cercenado E Dauden E Blazquez JLadron de Guevara C Gomez J de la Torre JQuarterly 45 267ndash275

Desjeux P (1998) Leishmania and HIV in Gridlock Barros C Altes J Serra T amp Alvar J (1995)Variability of Leishmania (Leishmania) infantumDocument WHOCTDLEISH989 Geneva WHO

Desjeux P amp Alvar J (2003) LeishmaniaHIV among stocks from immunocompromised immuno-competent patients and dogs in Spain FEMSco-infections epidemiology in Europe Annals of

Tropical Medicine and Parasitology 97 (Suppl 1) Microbiology Letters 131 197ndash204Killick-Kendrick R (1990) Phlebotomine vectors of3ndash15

Fillola G Corberand J X Laharrague P F the leishmaniases a review Medical and VeterinaryEntomology 4 1ndash24Levenes H Massip P amp Recco P (1992)

Peripheral intramonocytic leishmanias in an AIDS Kubar J Quaranta J F Aufeuvre J P Marty PLelievre A amp Le Fichoux Y (1997) Transmissionpatient Journal of Clinical Microbiology 30 3284ndash3285

Freedman D O MacLean J D amp Viloria J B of Leishmania infantum by blood donnors NatureMedicine 3 368(1987) A case of laboratory acquired Leishmania

donovani infection evidence for primary lymphatic Kubar J Marty P Lelievre A Quaranta J FStaccini P Caroli-Bosc C amp Le Fichoux Ydissemination Transactions of the Royal Society of

Tropical Medicine and Hygiene 81 118ndash119 (1998) Visceral leishmaniosis in HIV-positivepatients primary infection reactivation and latentGradoni L Gramiccia M Leger N Pesson B

Madulo-Leblond G Killick-Kendrick R Killick- infection Impact of the CD4+ T-lymphocyte countsAIDS 12 2147ndash2153Kendrick M amp Walton B C (1991) Isoenzyme

characterization of Leishmania from man dog and Le Fichoux Y Quaranta J F Aufeuvre J PLelievre A Marty P SuYa I Rousseau D ampsand ies in the Maltese islands Transactions of the

Royal Society of Tropical Medicine and Hygiene 85 Kubar J (1999) Occurrence of Leishmania infantumparasitemia in asymptomatic blood donors living in217ndash219

Gradoni L Pizzuti R Di Martino L Gramiccia M an area of endemicity in southern France Journal ofClinical Microbiology 37 1953ndash1957Pempinello R Gaeta G B Ferrara M Scotti S

amp Altieri S (1993) The epidemiology and surveil- Lopez-Velez R Laguna F Alvar J Perez-MolinaJ A Molina R Martinez P amp Villarrubia Jlance of visceral leishmaniasis in the Campania region

of Italy The value of zymodeme typing Epidemiology (1995) Parasitic culture of buVy coat for diagnosisof visceral leishmaniasis in human immunode ciencyand Infection 111 297ndash306

Gradoni L Scalone A Gramiccia M amp Troiani M virus-infected patients Journal of Clinical Micro-biology 33 937ndash939(1996) Epidemiological surveillance of leishmaniasis

44 MOLINA ET AL

Luz K G da Silva V O Gomes E M Quilici M amp Kaplanski S (1991) Evidence of sub-jets sensitized to Leishmania infantum on the FrenchMachado F C Araujo M A Fonseca H E

Freire T C drsquoAlmeida J B Palatnik M amp Mediterranean coast diVerences in gamma interferonproduction between this population and visceralPalatnik-de Sousa C B (1997) Prevalence of anti-

Leishmania donovani antibody among Brazilian blood leishmaniasis patients Parasite Immunology 13531ndash536donors and multiply transfused hemodialysis patients

American Journal of Tropical Medicine and Hygiene Molina R (1991) Laboratory adaptation of an auto-chthonous colony of Phlebotomus perniciosus Newstead57 168ndash171

Lysenko A J amp Beljaev A E (1987) Quantitative 1911 (Diptera Psychodidae) Research and Reviewsin Parasitology 51 87ndash89approaches to epidemiology In The Leishmaniases in

Biology and Medicine Vol 1 eds Peters W amp Killick- Molina R amp Alvar J (1996) A simple protocol forthe indirect xenodiagnosis of Leishmania infantum inKendrick R pp 263ndash290 London Academic Press

Ma D D Concannon A J amp Hayes J (1979) Fatal the blood of HIV-infected patients Annals of TropicalMedicine and Parasitology 90 639ndash640leishmaniasis in renal-transport patient Lancet ii

311ndash312 Molina R Lopez-Velez R Gutierrez-Solar BJimenez M I amp Alvar J (1992) Isolation ofMartotilde n-Sanchez J Guilvard E Acedo-Sanchez C

Wolf-Echeverri M Sanchiz-Mar otilde n M C amp Leishmania infantum from the blood of a patient withAIDS using sand ies Transactions of the Royal SocietyMorillas-Marquez F (1994) Phlebotomus perniciosus

Newstead 1911 infection by various zymodemes of of Tropical Medicine and Hygiene 86 516Molina R Canavate C Cercenado E Laguna Fthe Leishmania infantum complex in the Granada

province (southern Spain) International Journal for Lopez-Velez R amp Alvar J (1994) Indirect xeno-diagnosis of visceral leishmaniasis in 10 HIV-infectedParasitology 24 405ndash408

Martotilde n-Sanchez J Morillas-Marquez F Acedo- patients using colonized Phlebotomus perniciosusAIDS 8 277ndash279Sanchez C amp Sanchiz-Marotilde n M C (1995) The

variability of the etiological agent of leishmaniasis in Molina R Lohse J M amp Alvar J (1998) Infectividadde individuos inmunocompetentes e inmumo-the northndasheast of the Almer otilde a region southndasheast

Spain Systematic Parasitology 30 233ndash238 deprimidos parasitados por Leishmania infantum InI Congreso de la Sociedad Espanola de Medicina TropicalMartotilde n-Sanchez J Ru otilde z-Martotilde nez F Salinas-

Martotilde nez de Lecea J M Sanchez-Rabasco C y Salud Internacional (Chinchon Spain) p 111Madrid Unidad de Investigacion en EnfermedadesAcedo-Sanchez C Sanchiz-Mar otilde n M C Delgado-

Florencio V amp Morillas-Marquez F (1996) Tropicales y Salud Internacional Instituto de SaludlsquoCarlos IIIrsquoLeishmania infantum Nicolle 1908 from southern

Spain Characterization of the strains from human Molina R Lohse J M Pulido F Laguna FLopez-Velez R amp Alvar J (1999) Infection of sandvisceral and cutaneous leishmaniasis and from sand-

ies with a numerical analysis of the isoenzymatic ies by humans coinfected with Leishmania infantumand human immunode ciency virus Americandata Systematic Parasitology 33 177ndash182

Martotilde nez J E Arias A L Escobar M A amp Journal of Tropical Medicine and Hygiene 60 51ndash53Morillas F Sanchez Rabasco F Ocana J Martotilde nSaravia N G (1992) Haemoculture of Leishmania

( Viannia) braziliensis from two cases of mucosal Sanchez J Ocana Wihelmi J Acedo C amp Sanchotilde zMarotilde n M C (1996) Leishmaniosis in the focus ofleishmaniasis re-examination of haematogenous dis-

semination Transactions of the Royal Society of the Axarqu otilde a region Malaga province southernSpain a survey of the human dog and vectorTropical Medicine and Hygiene 86 392ndash394

Martotilde nez P de la Vega E Laguna F Soriano V Parasitology Research 82 569ndash570Otero A C da Silva V O Luz K G Palatnik MPuente S Moreno V Sentchordi M J Garcotilde a-

Aguado C amp Gonzalez-Lahoz J (1993) Diagnosis Pirmez C Fernandes O amp Palatnik de Sousa C B(2000) Occurrence of Leishmania donovani DNAof visceral leishmaniasis in HIV-infected individuals

using peripheral blood smears AIDS 7 227ndash230 in donated blood from seroreactive Brazilian blooddonors American Journal of Tropical Medicine andMarty P Le Fichoux Y Giordana D amp Brugnetti A

(1992) Leishmanin reaction in the human popu- Hygiene 62 128ndash131Owens S D Oakley D A Marryott Klation of a highly endemic focus of canine leishmaniasis

in Alpes-Maritimes France Transactions of the Royal Hatchett W Walton R Nolan T J Newton ASteurer F Schantz P amp Giger U (2001)Society of Tropical Medicine and Hygiene 86 249ndash250

Medrano F J Jimenez-Mejotilde as E Calderon E Transmission of visceral leishmaniasis through bloodtransfusions from infected English foxhounds toRegordan C amp Leal M (1993) An easy and quick

method for the dignosis of visceral leishmaniasis in anemic dogs Journal of the American VeterinaryMedical Association 219 1076ndash1083HIV-1-infected individuals AIDS 13 1399

Meller-Melloul C Farnarier C Dunan S Palatnik de Sousa C B Paraguai de Souza EGomes E M Soares-Machado F C Luz K GFaugere B Franck J Mary C Bongrand P


amp Borojevic R (1996) Transmission of visceral Rezza G (1998) Determinants of progression to AIDSin HIV-infected individuals an update from theleishmaniasis by blood transfusion in hamsters

Brazilian Journal of Medical and Biological Research Italian Seroconversion Study Journal of AcquiredImmune De ciency Syndromes and Human Retro-29 1311ndash1315

Pampiglione S Manson-Bahr P E C La-Placa M virology 17 (Suppl 1) S13ndashS16Rioux J A Guilvard E Gallego J Moreno GBorgatti M A amp Musumeci M (1975) Studies on

Mediterranean leishmaniasis 3 The leishmanin skin Pratlong F Portus M Rispail P Gallego M ampBastien P (1986) Phlebotomus ariasi Tonnoir 1921test in kala-azar Transactions of the Royal Society of

Tropical Medicine and Hygiene 69 60ndash68 et Phlebotomus perniciosus Newstead 1911 vecteursdu complexe Leishmania infantum dans un memePampiglione S Manson-Bahr P E C La-Placa M

Borgatti M A amp Micheloni F (1976) Studies on foyer Infestations par deux zymodemes syntopiquesA propos drsquoune enquete en Catalogne (Espagne)Mediterranean leishmaniasis IV The leishmanin

skin test in cutaneous leishmaniasis Transactions of In Leishmania Taxonomie et Phylogenese ApplicationsEco-epidemiologique (Colloques Internationales duthe Royal Society of Tropical Medicine and Hygiene

70 62ndash65 CNRSINSERM) ed Rioux J A pp 439ndash444Montpellier Institut Mediterraneen drsquoEtudes Epi-Pezzotti P Scalia Tomba G amp Rezza G (1995)

Incidenza e Prevalenza delle Infezioni da HIV in Italia demiologiques et EcologiquesRosenthal E Marty P Poizot-Martin I Reynes Je Previsione a Breve e Medio Termine (Rapporti

ISTISAN 9541) Rome Istituto Superiore di Sanita Pratlong F Lafeuillade A Jaubert D Boulat ODereure J Gambarelli F Gastaut J APineda J A Gallardo J A Mac otilde as J Delgado J

Regordan C Morillas F Relimpio F Martotilde n- Dujardin P Dellamonica P amp Cassuto J (1995)Visceral leishmaniasis and HIV-1 co-infection inSanchez J Sanchez-Quijano A Leal M amp LissenE

(1998) Prevalence of and factors associated with southern France Transactions of the Royal Society ofTropical Medicine and Hygiene 89 159ndash162visceral leishmaniasis in human immunode ciency

virus type 1-infected patients in southern Spain Singh S Chaudhry V P amp Wali J P (1996)Transfusion-transmitted kala-azar in India Trans-Journal of Clinical Microbiology 36 2419ndash2422

Pratlong F Dedet J P Marty P Portus M fusion 36 848ndash849Tesh R B (1995) Control of zoonotic visceralDeniau M Dereure J Abranches P

Reynes J Martini A Lefebvre M amp Rioux J A leishmaniasis is it time to change strategies AmericanJournal of Tropical Medicine and Hygiene 52 287ndash292(1995) Leishmaniandashhuman immunode ciency virus

coinfection in the Mediterranean basin isoenzymatic Theodor O (1964) Leishmaniases In Zoonoses edvan der Hoeden J pp 475ndash493 Amsterdam Elseviercharacterization of 100 isolates of the Leishmania

infantum complex Journal of Infectious Diseases 172 World Health Organization (1998) Report on the ThirdConsultative Meeting on LeishmaniaHIV Co-infections323ndash326

Rab M A Hassan M Bux D Mahmood M T held in Mahon (Spain) Document WHOCTDLEISH Geneva WHOamp Evans D A (1992) The isolation and cultivation

of Leishmania infantum from apparently normal World Health Organization (1999) LeishmaniaHIVco-infection southndashwestern Europe 1990ndash1998 Retro-skin of visceral leishmaniasis patients in northern

Pakistan Transactions of the Royal Society of Tropical spective analysis of 965 cases Weekly EpidemiologicalRecord 74 365ndash375Medicine and Hygiene 86 620ndash621

30 MOLINA ET AL

transmitted from human to human by the and Spain give positive results in leishmaninskin tests (Pampiglione et al 1975 1976bite of a species of sand y that is strictly

peridomestic P argentipes Gramiccia et al 1990 Meller-Melloul et al1991 Marty et al 1992 Arbaji et al 1993Since the global epidemic of HIV infection

began there has been a steady rise in the Acedo-Sanchez et al 1996 Alvar et al1996 Morillas et al 1996) many detectedannual number of reported new cases of VL

associated with the virus LeishmaniaHIV cases of LeishmaniaHIV co-infection in thesecountries are probably the result of latentco-infection is emerging as a new and worry-

ing disease particularly in southndashwestern Leishmania infections that have been activatedor re-activated as the later HIV infectionEurope (Desjeux 1998) In Spain mdash the

country reporting the highest incidence of causes immunosuppression Many now thinkthat Leishmania spp should be consideredthe co-infection mdash 68 of the cases are

intravenous-drug users (Desjeux and Alvar as opportunistic pathogens in HIV-infectedpatients (Alvar et al 1992) Even in the2003) In France Italy Portugal and Spain

the outbreak of HIV infection has changed absence of HIV Le infantum infectionsmay be re-activated following immunologicalthe epidemiology of VL from a disease pre-

dominantly found in children to one more distress (Ma et al 1979 Kubar et al 1998)If the natural history of VL in HIV-common in adults (WHO 1999) HIV co-

infection may re-activate latent leishmanial positive individuals is to be fully understoodlong-term longitudinal immunological andinfections increase the level of transmission

of Leishmania (particularly humanndashhuman parasitological investigations on cohorts ofindividuals at risk of both HIV and Leishmaniatransmission) by phlebotomine sand ies and

faciliate arti cial transmission of Leishmania infection will have to be conducted As theincidence of leishmaniasis in the areas mostvia the sharing of contaminated syringes and

needles from one intravenous-drug user aVected by the co-infection is relatively lowthe cohorts studied will have to be large The(IVDU) to another (see below)relevant epidemiological data already availablemay be suYcient however to estimate the

NEWLY ACQUIRED VERSUS probabilities that clinical VL in a HIV-positiveRE-ACTIVATED VISCERAL individual is (1) the result of a newly acquired

LEISHMANIASIS THE ITALIAN leishmanial infection or (2) the result ofEXAMPLE a re-activated latent infection In 1998

L Gradoni and A Scalone attempted toestimate these two probabilities in SicilyIt is estimated that only one in every ve to

10 immunocompetent individuals who have by assimilating and updating the relevantepidemiological-surveillance data and thenbeen infected with Le donovani or Le infantum

ever develops clinical VL (Desjeux 1992 using the resultant data-set in an epidemio-logical model (unpubl obs) Their analysisBadaro et al 1996) Since a T-lymphocyte-

mediated immune response is necessary to of the natural history of the LeishmaniaHIVco-infection is described below in extensocontrol Leishmania infection however those

residents of endemic areas who have beenmade immunode cient as the result of HIVinfection are far more prone to VL than their Surveillance Methodology

In 1989 a programme based on the activeimmunocompetent neighbours Strains ofLeishmania that are usually non-pathogenic detection of LeishmaniaHIV co-infection was

implemented in collaboration with some 148in humans and even lower trypanosomatids(Chicharro and Alvar 2003) may all cause infectious-disease units sited in universities

and hospitals throughout Italy (Gradoni et alillness in HIV-positive individuals As manyasymptomatic individuals in Italy France 1996) The units selected from the 260

















32 MOLINA ET AL























34 MOLINA ET AL
































36 MOLINA ET AL












































38 MOLINA ET AL

TA

BL

E1

C

linic

alan

dpa

rasi

tolo

gica

lda

tafo

r10

case

sof

Lei

shm

ania

infa

ntu

mH

IVco

-inf

ecti

onin

vest

igat

edby

indi

rect

xeno

diag

nosi

s

Cas

e

12

34

56

78

91

0

Sex

MM

MF

MM

MM

MM

Age

(yea

rs)

36

333

525

382

922

2828

37

Exp

osu

regr

oup

for

HIV

infe

ctio

nIV

DU

IVD

UIV

DU

IVD

UH

SIV

DU

IVD

UIV

DU

IVD

UIV

DU

Ass

ocia

ted

infe

ctio

n(s

)P

TB

O

CC

MV

PC

PE

CP

TB

E

CO

CE

PT

BN

oN

oP

CP

Fev

er

Yes

No

Yes

Yes

Yes

Yes

Yes

Yes

Yes

Yes

Spl

enom

egal

yY

esN

oY

esY

esY

esY

esY

esY

esY

esN

oL

euco

cyte

cou

nt

(leu

cocy

tes

ml)

270

02

400

NA

250

043

0036

0024

00

1600

190

059

00C

D4

+co

un

t(C

D4

+ce

llsm

l)4

010

68

5648

51

144

201

48N

DH

aem

oglo

bin

(gl

)8

48

3N

A7

28

49

10

110

65

74

Pla

tele

tco

un

t(p

late

lets

nl)

71

164

NA

8475

98

7317

91

311

70A

mas

tigo

tes

inb

lood

smea

rN

oY

esN

oY

esY

esY

esN

oN

oN

oY

esA

mas

tigo

tes

insm

ear

ofbo

ne-

mar

row

aspi

rate

N

oY

esY

esY

esY

esY

esN

oN

oN

oY

esS

erol

ogy

(IF

AT

titr

e)1

160

lt1

40

18

0lt

140

lt1

40lt

140

116

01

320

140

lt1

40B

one-

mar

row

cult

ure

++

++

++

shy+

shy+

Per

iphe

ral-

bloo

d-m

onoc

yte

cult

ure

shy+

++

shy+

++

shy+

Tre

atm

ent

for

leis

hm

ania

sis

(day

s)2

00

00

0R

elap

seR

elap

seR

elap

se0

Res

ult

ofin

dire

ctxe

nod

iagn

osis

++

++

++

++

++

Blo

odst

orag

eat

4degC

pri

orto

xen

odia

gnos

is(h

)6

62

46

366

2448

722

4

ofb

lood

fed

san

d

ies

fou

ndin

fect

ed22

837

514

262

79

29

48

38

010

713

853

3

M

Mal

eF

fe

mal

eIV

DU

in

trav

enou

s-dr

ug

use

rH

S

hete

rose

xual

P

TB

p

ulm

onar

ytu

berc

ulos

is

OC

or

alca

nd

idia

sis

CM

V

cyto

meg

alov

iru

sin

fect

ion

P

CP

P

neum

ocys

tisca

rini

iE

PT

B

extr

apu

lmon

ary

tub

ercu

losi

sN

A

not

avai

lab

le

ND

n

otd

one

























40 MOLINA ET AL



Case

1 2 3 4 5 6





















42 MOLINA ET AL





















































44 MOLINA ET AL



































































32 MOLINA ET AL























34 MOLINA ET AL
































36 MOLINA ET AL












































38 MOLINA ET AL

TA

BL

E1

C

linic

alan

dpa

rasi

tolo

gica

lda

tafo

r10

case

sof

Lei

shm

ania

infa

ntu

mH

IVco

-inf

ecti

onin

vest

igat

edby

indi

rect

xeno

diag

nosi

s

Cas

e

12

34

56

78

91

0

Sex

MM

MF

MM

MM

MM

Age

(yea

rs)

36

333

525

382

922

2828

37

Exp

osu

regr

oup

for

HIV

infe

ctio

nIV

DU

IVD

UIV

DU

IVD

UH

SIV

DU

IVD

UIV

DU

IVD

UIV

DU

Ass

ocia

ted

infe

ctio

n(s

)P

TB

O

CC

MV

PC

PE

CP

TB

E

CO

CE

PT

BN

oN

oP

CP

Fev

er

Yes

No

Yes

Yes

Yes

Yes

Yes

Yes

Yes

Yes

Spl

enom

egal

yY

esN

oY

esY

esY

esY

esY

esY

esY

esN

oL

euco

cyte

cou

nt

(leu

cocy

tes

ml)

270

02

400

NA

250

043

0036

0024

00

1600

190

059

00C

D4

+co

un

t(C

D4

+ce

llsm

l)4

010

68

5648

51

144

201

48N

DH

aem

oglo

bin

(gl

)8

48

3N

A7

28

49

10

110

65

74

Pla

tele

tco

un

t(p

late

lets

nl)

71

164

NA

8475

98

7317

91

311

70A

mas

tigo

tes

inb

lood

smea

rN

oY

esN

oY

esY

esY

esN

oN

oN

oY

esA

mas

tigo

tes

insm

ear

ofbo

ne-

mar

row

aspi

rate

N

oY

esY

esY

esY

esY

esN

oN

oN

oY

esS

erol

ogy

(IF

AT

titr

e)1

160

lt1

40

18

0lt

140

lt1

40lt

140

116

01

320

140

lt1

40B

one-

mar

row

cult

ure

++

++

++

shy+

shy+

Per

iphe

ral-

bloo

d-m

onoc

yte

cult

ure

shy+

++

shy+

++

shy+

Tre

atm

ent

for

leis

hm

ania

sis

(day

s)2

00

00

0R

elap

seR

elap

seR

elap

se0

Res

ult

ofin

dire

ctxe

nod

iagn

osis

++

++

++

++

++

Blo

odst

orag

eat

4degC

pri

orto

xen

odia

gnos

is(h

)6

62

46

366

2448

722

4

ofb

lood

fed

san

d

ies

fou

ndin

fect

ed22

837

514

262

79

29

48

38

010

713

853

3

M

Mal

eF

fe

mal

eIV

DU

in

trav

enou

s-dr

ug

use

rH

S

hete

rose

xual

P

TB

p

ulm

onar

ytu

berc

ulos

is

OC

or

alca

nd

idia

sis

CM

V

cyto

meg

alov

iru

sin

fect

ion

P

CP

P

neum

ocys

tisca

rini

iE

PT

B

extr

apu

lmon

ary

tub

ercu

losi

sN

A

not

avai

lab

le

ND

n

otd

one

























40 MOLINA ET AL



Case

1 2 3 4 5 6





















42 MOLINA ET AL





















































44 MOLINA ET AL



















































32 MOLINA ET AL























34 MOLINA ET AL
































36 MOLINA ET AL












































38 MOLINA ET AL

TA

BL

E1

C

linic

alan

dpa

rasi

tolo

gica

lda

tafo

r10

case

sof

Lei

shm

ania

infa

ntu

mH

IVco

-inf

ecti

onin

vest

igat

edby

indi

rect

xeno

diag

nosi

s

Cas

e

12

34

56

78

91

0

Sex

MM

MF

MM

MM

MM

Age

(yea

rs)

36

333

525

382

922

2828

37

Exp

osu

regr

oup

for

HIV

infe

ctio

nIV

DU

IVD

UIV

DU

IVD

UH

SIV

DU

IVD

UIV

DU

IVD

UIV

DU

Ass

ocia

ted

infe

ctio

n(s

)P

TB

O

CC

MV

PC

PE

CP

TB

E

CO

CE

PT

BN

oN

oP

CP

Fev

er

Yes

No

Yes

Yes

Yes

Yes

Yes

Yes

Yes

Yes

Spl

enom

egal

yY

esN

oY

esY

esY

esY

esY

esY

esY

esN

oL

euco

cyte

cou

nt

(leu

cocy

tes

ml)

270

02

400

NA

250

043

0036

0024

00

1600

190

059

00C

D4

+co

un

t(C

D4

+ce

llsm

l)4

010

68

5648

51

144

201

48N

DH

aem

oglo

bin

(gl

)8

48

3N

A7

28

49

10

110

65

74

Pla

tele

tco

un

t(p

late

lets

nl)

71

164

NA

8475

98

7317

91

311

70A

mas

tigo

tes

inb

lood

smea

rN

oY

esN

oY

esY

esY

esN

oN

oN

oY

esA

mas

tigo

tes

insm

ear

ofbo

ne-

mar

row

aspi

rate

N

oY

esY

esY

esY

esY

esN

oN

oN

oY

esS

erol

ogy

(IF

AT

titr

e)1

160

lt1

40

18

0lt

140

lt1

40lt

140

116

01

320

140

lt1

40B

one-

mar

row

cult

ure

++

++

++

shy+

shy+

Per

iphe

ral-

bloo

d-m

onoc

yte

cult

ure

shy+

++

shy+

++

shy+

Tre

atm

ent

for

leis

hm

ania

sis

(day

s)2

00

00

0R

elap

seR

elap

seR

elap

se0

Res

ult

ofin

dire

ctxe

nod

iagn

osis

++

++

++

++

++

Blo

odst

orag

eat

4degC

pri

orto

xen

odia

gnos

is(h

)6

62

46

366

2448

722

4

ofb

lood

fed

san

d

ies

fou

ndin

fect

ed22

837

514

262

79

29

48

38

010

713

853

3

M

Mal

eF

fe

mal

eIV

DU

in

trav

enou

s-dr

ug

use

rH

S

hete

rose

xual

P

TB

p

ulm

onar

ytu

berc

ulos

is

OC

or

alca

nd

idia

sis

CM

V

cyto

meg

alov

iru

sin

fect

ion

P

CP

P

neum

ocys

tisca

rini

iE

PT

B

extr

apu

lmon

ary

tub

ercu

losi

sN

A

not

avai

lab

le

ND

n

otd

one

























40 MOLINA ET AL



Case

1 2 3 4 5 6





















42 MOLINA ET AL





















































44 MOLINA ET AL





























































34 MOLINA ET AL
































36 MOLINA ET AL












































38 MOLINA ET AL

TA

BL

E1

C

linic

alan

dpa

rasi

tolo

gica

lda

tafo

r10

case

sof

Lei

shm

ania

infa

ntu

mH

IVco

-inf

ecti

onin

vest

igat

edby

indi

rect

xeno

diag

nosi

s

Cas

e

12

34

56

78

91

0

Sex

MM

MF

MM

MM

MM

Age

(yea

rs)

36

333

525

382

922

2828

37

Exp

osu

regr

oup

for

HIV

infe

ctio

nIV

DU

IVD

UIV

DU

IVD

UH

SIV

DU

IVD

UIV

DU

IVD

UIV

DU

Ass

ocia

ted

infe

ctio

n(s

)P

TB

O

CC

MV

PC

PE

CP

TB

E

CO

CE

PT

BN

oN

oP

CP

Fev

er

Yes

No

Yes

Yes

Yes

Yes

Yes

Yes

Yes

Yes

Spl

enom

egal

yY

esN

oY

esY

esY

esY

esY

esY

esY

esN

oL

euco

cyte

cou

nt

(leu

cocy

tes

ml)

270

02

400

NA

250

043

0036

0024

00

1600

190

059

00C

D4

+co

un

t(C

D4

+ce

llsm

l)4

010

68

5648

51

144

201

48N

DH

aem

oglo

bin

(gl

)8

48

3N

A7

28

49

10

110

65

74

Pla

tele

tco

un

t(p

late

lets

nl)

71

164

NA

8475

98

7317

91

311

70A

mas

tigo

tes

inb

lood

smea

rN

oY

esN

oY

esY

esY

esN

oN

oN

oY

esA

mas

tigo

tes

insm

ear

ofbo

ne-

mar

row

aspi

rate

N

oY

esY

esY

esY

esY

esN

oN

oN

oY

esS

erol

ogy

(IF

AT

titr

e)1

160

lt1

40

18

0lt

140

lt1

40lt

140

116

01

320

140

lt1

40B

one-

mar

row

cult

ure

++

++

++

shy+

shy+

Per

iphe

ral-

bloo

d-m

onoc

yte

cult

ure

shy+

++

shy+

++

shy+

Tre

atm

ent

for

leis

hm

ania

sis

(day

s)2

00

00

0R

elap

seR

elap

seR

elap

se0

Res

ult

ofin

dire

ctxe

nod

iagn

osis

++

++

++

++

++

Blo

odst

orag

eat

4degC

pri

orto

xen

odia

gnos

is(h

)6

62

46

366

2448

722

4

ofb

lood

fed

san

d

ies

fou

ndin

fect

ed22

837

514

262

79

29

48

38

010

713

853

3

M

Mal

eF

fe

mal

eIV

DU

in

trav

enou

s-dr

ug

use

rH

S

hete

rose

xual

P

TB

p

ulm

onar

ytu

berc

ulos

is

OC

or

alca

nd

idia

sis

CM

V

cyto

meg

alov

iru

sin

fect

ion

P

CP

P

neum

ocys

tisca

rini

iE

PT

B

extr

apu

lmon

ary

tub

ercu

losi

sN

A

not

avai

lab

le

ND

n

otd

one

























40 MOLINA ET AL



Case

1 2 3 4 5 6





















42 MOLINA ET AL





















































44 MOLINA ET AL



















































34 MOLINA ET AL
































36 MOLINA ET AL












































38 MOLINA ET AL

TA

BL

E1

C

linic

alan

dpa

rasi

tolo

gica

lda

tafo

r10

case

sof

Lei

shm

ania

infa

ntu

mH

IVco

-inf

ecti

onin

vest

igat

edby

indi

rect

xeno

diag

nosi

s

Cas

e

12

34

56

78

91

0

Sex

MM

MF

MM

MM

MM

Age

(yea

rs)

36

333

525

382

922

2828

37

Exp

osu

regr

oup

for

HIV

infe

ctio

nIV

DU

IVD

UIV

DU

IVD

UH

SIV

DU

IVD

UIV

DU

IVD

UIV

DU

Ass

ocia

ted

infe

ctio

n(s

)P

TB

O

CC

MV

PC

PE

CP

TB

E

CO

CE

PT

BN

oN

oP

CP

Fev

er

Yes

No

Yes

Yes

Yes

Yes

Yes

Yes

Yes

Yes

Spl

enom

egal

yY

esN

oY

esY

esY

esY

esY

esY

esY

esN

oL

euco

cyte

cou

nt

(leu

cocy

tes

ml)

270

02

400

NA

250

043

0036

0024

00

1600

190

059

00C

D4

+co

un

t(C

D4

+ce

llsm

l)4

010

68

5648

51

144

201

48N

DH

aem

oglo

bin

(gl

)8

48

3N

A7

28

49

10

110

65

74

Pla

tele

tco

un

t(p

late

lets

nl)

71

164

NA

8475

98

7317

91

311

70A

mas

tigo

tes

inb

lood

smea

rN

oY

esN

oY

esY

esY

esN

oN

oN

oY

esA

mas

tigo

tes

insm

ear

ofbo

ne-

mar

row

aspi

rate

N

oY

esY

esY

esY

esY

esN

oN

oN

oY

esS

erol

ogy

(IF

AT

titr

e)1

160

lt1

40

18

0lt

140

lt1

40lt

140

116

01

320

140

lt1

40B

one-

mar

row

cult

ure

++

++

++

shy+

shy+

Per

iphe

ral-

bloo

d-m

onoc

yte

cult

ure

shy+

++

shy+

++

shy+

Tre

atm

ent

for

leis

hm

ania

sis

(day

s)2

00

00

0R

elap

seR

elap

seR

elap

se0

Res

ult

ofin

dire

ctxe

nod

iagn

osis

++

++

++

++

++

Blo

odst

orag

eat

4degC

pri

orto

xen

odia

gnos

is(h

)6

62

46

366

2448

722

4

ofb

lood

fed

san

d

ies

fou

ndin

fect

ed22

837

514

262

79

29

48

38

010

713

853

3

M

Mal

eF

fe

mal

eIV

DU

in

trav

enou

s-dr

ug

use

rH

S

hete

rose

xual

P

TB

p

ulm

onar

ytu

berc

ulos

is

OC

or

alca

nd

idia

sis

CM

V

cyto

meg

alov

iru

sin

fect

ion

P

CP

P

neum

ocys

tisca

rini

iE

PT

B

extr

apu

lmon

ary

tub

ercu

losi

sN

A

not

avai

lab

le

ND

n

otd

one

























40 MOLINA ET AL



Case

1 2 3 4 5 6





















42 MOLINA ET AL





















































44 MOLINA ET AL





































































36 MOLINA ET AL












































38 MOLINA ET AL

TA

BL

E1

C

linic

alan

dpa

rasi

tolo

gica

lda

tafo

r10

case

sof

Lei

shm

ania

infa

ntu

mH

IVco

-inf

ecti

onin

vest

igat

edby

indi

rect

xeno

diag

nosi

s

Cas

e

12

34

56

78

91

0

Sex

MM

MF

MM

MM

MM

Age

(yea

rs)

36

333

525

382

922

2828

37

Exp

osu

regr

oup

for

HIV

infe

ctio

nIV

DU

IVD

UIV

DU

IVD

UH

SIV

DU

IVD

UIV

DU

IVD

UIV

DU

Ass

ocia

ted

infe

ctio

n(s

)P

TB

O

CC

MV

PC

PE

CP

TB

E

CO

CE

PT

BN

oN

oP

CP

Fev

er

Yes

No

Yes

Yes

Yes

Yes

Yes

Yes

Yes

Yes

Spl

enom

egal

yY

esN

oY

esY

esY

esY

esY

esY

esY

esN

oL

euco

cyte

cou

nt

(leu

cocy

tes

ml)

270

02

400

NA

250

043

0036

0024

00

1600

190

059

00C

D4

+co

un

t(C

D4

+ce

llsm

l)4

010

68

5648

51

144

201

48N

DH

aem

oglo

bin

(gl

)8

48

3N

A7

28

49

10

110

65

74

Pla

tele

tco

un

t(p

late

lets

nl)

71

164

NA

8475

98

7317

91

311

70A

mas

tigo

tes

inb

lood

smea

rN

oY

esN

oY

esY

esY

esN

oN

oN

oY

esA

mas

tigo

tes

insm

ear

ofbo

ne-

mar

row

aspi

rate

N

oY

esY

esY

esY

esY

esN

oN

oN

oY

esS

erol

ogy

(IF

AT

titr

e)1

160

lt1

40

18

0lt

140

lt1

40lt

140

116

01

320

140

lt1

40B

one-

mar

row

cult

ure

++

++

++

shy+

shy+

Per

iphe

ral-

bloo

d-m

onoc

yte

cult

ure

shy+

++

shy+

++

shy+

Tre

atm

ent

for

leis

hm

ania

sis

(day

s)2

00

00

0R

elap

seR

elap

seR

elap

se0

Res

ult

ofin

dire

ctxe

nod

iagn

osis

++

++

++

++

++

Blo

odst

orag

eat

4degC

pri

orto

xen

odia

gnos

is(h

)6

62

46

366

2448

722

4

ofb

lood

fed

san

d

ies

fou

ndin

fect

ed22

837

514

262

79

29

48

38

010

713

853

3

M

Mal

eF

fe

mal

eIV

DU

in

trav

enou

s-dr

ug

use

rH

S

hete

rose

xual

P

TB

p

ulm

onar

ytu

berc

ulos

is

OC

or

alca

nd

idia

sis

CM

V

cyto

meg

alov

iru

sin

fect

ion

P

CP

P

neum

ocys

tisca

rini

iE

PT

B

extr

apu

lmon

ary

tub

ercu

losi

sN

A

not

avai

lab

le

ND

n

otd

one

























40 MOLINA ET AL



Case

1 2 3 4 5 6





















42 MOLINA ET AL





















































44 MOLINA ET AL



















































36 MOLINA ET AL












































38 MOLINA ET AL

TA

BL

E1

C

linic

alan

dpa

rasi

tolo

gica

lda

tafo

r10

case

sof

Lei

shm

ania

infa

ntu

mH

IVco

-inf

ecti

onin

vest

igat

edby

indi

rect

xeno

diag

nosi

s

Cas

e

12

34

56

78

91

0

Sex

MM

MF

MM

MM

MM

Age

(yea

rs)

36

333

525

382

922

2828

37

Exp

osu

regr

oup

for

HIV

infe

ctio

nIV

DU

IVD

UIV

DU

IVD

UH

SIV

DU

IVD

UIV

DU

IVD

UIV

DU

Ass

ocia

ted

infe

ctio

n(s

)P

TB

O

CC

MV

PC

PE

CP

TB

E

CO

CE

PT

BN

oN

oP

CP

Fev

er

Yes

No

Yes

Yes

Yes

Yes

Yes

Yes

Yes

Yes

Spl

enom

egal

yY

esN

oY

esY

esY

esY

esY

esY

esY

esN

oL

euco

cyte

cou

nt

(leu

cocy

tes

ml)

270

02

400

NA

250

043

0036

0024

00

1600

190

059

00C

D4

+co

un

t(C

D4

+ce

llsm

l)4

010

68

5648

51

144

201

48N

DH

aem

oglo

bin

(gl

)8

48

3N

A7

28

49

10

110

65

74

Pla

tele

tco

un

t(p

late

lets

nl)

71

164

NA

8475

98

7317

91

311

70A

mas

tigo

tes

inb

lood

smea

rN

oY

esN

oY

esY

esY

esN

oN

oN

oY

esA

mas

tigo

tes

insm

ear

ofbo

ne-

mar

row

aspi

rate

N

oY

esY

esY

esY

esY

esN

oN

oN

oY

esS

erol

ogy

(IF

AT

titr

e)1

160

lt1

40

18

0lt

140

lt1

40lt

140

116

01

320

140

lt1

40B

one-

mar

row

cult

ure

++

++

++

shy+

shy+

Per

iphe

ral-

bloo

d-m

onoc

yte

cult

ure

shy+

++

shy+

++

shy+

Tre

atm

ent

for

leis

hm

ania

sis

(day

s)2

00

00

0R

elap

seR

elap

seR

elap

se0

Res

ult

ofin

dire

ctxe

nod

iagn

osis

++

++

++

++

++

Blo

odst

orag

eat

4degC

pri

orto

xen

odia

gnos

is(h

)6

62

46

366

2448

722

4

ofb

lood

fed

san

d

ies

fou

ndin

fect

ed22

837

514

262

79

29

48

38

010

713

853

3

M

Mal

eF

fe

mal

eIV

DU

in

trav

enou

s-dr

ug

use

rH

S

hete

rose

xual

P

TB

p

ulm

onar

ytu

berc

ulos

is

OC

or

alca

nd

idia

sis

CM

V

cyto

meg

alov

iru

sin

fect

ion

P

CP

P

neum

ocys

tisca

rini

iE

PT

B

extr

apu

lmon

ary

tub

ercu

losi

sN

A

not

avai

lab

le

ND

n

otd

one

























40 MOLINA ET AL



Case

1 2 3 4 5 6





















42 MOLINA ET AL





















































44 MOLINA ET AL



























































38 MOLINA ET AL

TA

BL

E1

C

linic

alan

dpa

rasi

tolo

gica

lda

tafo

r10

case

sof

Lei

shm

ania

infa

ntu

mH

IVco

-inf

ecti

onin

vest

igat

edby

indi

rect

xeno

diag

nosi

s

Cas

e

12

34

56

78

91

0

Sex

MM

MF

MM

MM

MM

Age

(yea

rs)

36

333

525

382

922

2828

37

Exp

osu

regr

oup

for

HIV

infe

ctio

nIV

DU

IVD

UIV

DU

IVD

UH

SIV

DU

IVD

UIV

DU

IVD

UIV

DU

Ass

ocia

ted

infe

ctio

n(s

)P

TB

O

CC

MV

PC

PE

CP

TB

E

CO

CE

PT

BN

oN

oP

CP

Fev

er

Yes

No

Yes

Yes

Yes

Yes

Yes

Yes

Yes

Yes

Spl

enom

egal

yY

esN

oY

esY

esY

esY

esY

esY

esY

esN

oL

euco

cyte

cou

nt

(leu

cocy

tes

ml)

270

02

400

NA

250

043

0036

0024

00

1600

190

059

00C

D4

+co

un

t(C

D4

+ce

llsm

l)4

010

68

5648

51

144

201

48N

DH

aem

oglo

bin

(gl

)8

48

3N

A7

28

49

10

110

65

74

Pla

tele

tco

un

t(p

late

lets

nl)

71

164

NA

8475

98

7317

91

311

70A

mas

tigo

tes

inb

lood

smea

rN

oY

esN

oY

esY

esY

esN

oN

oN

oY

esA

mas

tigo

tes

insm

ear

ofbo

ne-

mar

row

aspi

rate

N

oY

esY

esY

esY

esY

esN

oN

oN

oY

esS

erol

ogy

(IF

AT

titr

e)1

160

lt1

40

18

0lt

140

lt1

40lt

140

116

01

320

140

lt1

40B

one-

mar

row

cult

ure

++

++

++

shy+

shy+

Per

iphe

ral-

bloo

d-m

onoc

yte

cult

ure

shy+

++

shy+

++

shy+

Tre

atm

ent

for

leis

hm

ania

sis

(day

s)2

00

00

0R

elap

seR

elap

seR

elap

se0

Res

ult

ofin

dire

ctxe

nod

iagn

osis

++

++

++

++

++

Blo

odst

orag

eat

4degC

pri

orto

xen

odia

gnos

is(h

)6

62

46

366

2448

722

4

ofb

lood

fed

san

d

ies

fou

ndin

fect

ed22

837

514

262

79

29

48

38

010

713

853

3

M

Mal

eF

fe

mal

eIV

DU

in

trav

enou

s-dr

ug

use

rH

S

hete

rose

xual

P

TB

p

ulm

onar

ytu

berc

ulos

is

OC

or

alca

nd

idia

sis

CM

V

cyto

meg

alov

iru

sin

fect

ion

P

CP

P

neum

ocys

tisca

rini

iE

PT

B

extr

apu

lmon

ary

tub

ercu

losi

sN

A

not

avai

lab

le

ND

n

otd

one

























40 MOLINA ET AL



Case

1 2 3 4 5 6





















42 MOLINA ET AL





















































44 MOLINA ET AL



















































38 MOLINA ET AL

TA

BL

E1

C

linic

alan

dpa

rasi

tolo

gica

lda

tafo

r10

case

sof

Lei

shm

ania

infa

ntu

mH

IVco

-inf

ecti

onin

vest

igat

edby

indi

rect

xeno

diag

nosi

s

Cas

e

12

34

56

78

91

0

Sex

MM

MF

MM

MM

MM

Age

(yea

rs)

36

333

525

382

922

2828

37

Exp

osu

regr

oup

for

HIV

infe

ctio

nIV

DU

IVD

UIV

DU

IVD

UH

SIV

DU

IVD

UIV

DU

IVD

UIV

DU

Ass

ocia

ted

infe

ctio

n(s

)P

TB

O

CC

MV

PC

PE

CP

TB

E

CO

CE

PT

BN

oN

oP

CP

Fev

er

Yes

No

Yes

Yes

Yes

Yes

Yes

Yes

Yes

Yes

Spl

enom

egal

yY

esN

oY

esY

esY

esY

esY

esY

esY

esN

oL

euco

cyte

cou

nt

(leu

cocy

tes

ml)

270

02

400

NA

250

043

0036

0024

00

1600

190

059

00C

D4

+co

un

t(C

D4

+ce

llsm

l)4

010

68

5648

51

144

201

48N

DH

aem

oglo

bin

(gl

)8

48

3N

A7

28

49

10

110

65

74

Pla

tele

tco

un

t(p

late

lets

nl)

71

164

NA

8475

98

7317

91

311

70A

mas

tigo

tes

inb

lood

smea

rN

oY

esN

oY

esY

esY

esN

oN

oN

oY

esA

mas

tigo

tes

insm

ear

ofbo

ne-

mar

row

aspi

rate

N

oY

esY

esY

esY

esY

esN

oN

oN

oY

esS

erol

ogy

(IF

AT

titr

e)1

160

lt1

40

18

0lt

140

lt1

40lt

140

116

01

320

140

lt1

40B

one-

mar

row

cult

ure

++

++

++

shy+

shy+

Per

iphe

ral-

bloo

d-m

onoc

yte

cult

ure

shy+

++

shy+

++

shy+

Tre

atm

ent

for

leis

hm

ania

sis

(day

s)2

00

00

0R

elap

seR

elap

seR

elap

se0

Res

ult

ofin

dire

ctxe

nod

iagn

osis

++

++

++

++

++

Blo

odst

orag

eat

4degC

pri

orto

xen

odia

gnos

is(h

)6

62

46

366

2448

722

4

ofb

lood

fed

san

d

ies

fou

ndin

fect

ed22

837

514

262

79

29

48

38

010

713

853

3

M

Mal

eF

fe

mal

eIV

DU

in

trav

enou

s-dr

ug

use

rH

S

hete

rose

xual

P

TB

p

ulm

onar

ytu

berc

ulos

is

OC

or

alca

nd

idia

sis

CM

V

cyto

meg

alov

iru

sin

fect

ion

P

CP

P

neum

ocys

tisca

rini

iE

PT

B

extr

apu

lmon

ary

tub

ercu

losi

sN

A

not

avai

lab

le

ND

n

otd

one

























40 MOLINA ET AL



Case

1 2 3 4 5 6





















42 MOLINA ET AL





















































44 MOLINA ET AL











































































40 MOLINA ET AL



Case

1 2 3 4 5 6





















42 MOLINA ET AL





















































44 MOLINA ET AL



















































40 MOLINA ET AL



Case

1 2 3 4 5 6





















42 MOLINA ET AL





















































44 MOLINA ET AL
































































42 MOLINA ET AL





















































44 MOLINA ET AL



















































42 MOLINA ET AL





















































44 MOLINA ET AL

















































































44 MOLINA ET AL



















































44 MOLINA ET AL








































































Documents

HIV and the transmission of Leishmania - WHO...NEWLY ACQUIRED VERSUS probabilities thatclinicalVLinaHIV-positive RE-ACTIVATED VISCERAL individual is (1) the result of a newly acquired