Quantitative PCR for parasitemia monitoring and preemptive therapy in patients with Chagas disease and autoimmune rheumatic disorders undergoing biologic treatment: a case series

TL;DR

This study shows that monitoring parasitemia with qPCR can help prevent Chagas disease reactivation in patients with autoimmune disorders on immunosuppressive therapy.

Contribution

The study introduces the use of quantitative PCR for preemptive therapy in Chagas disease reactivation risk.

Findings

Five out of six patients showed parasitemia, with two having high parasitemia levels.

Benznidazole treatment led to a significant decrease in parasitemia in one patient.

No Chagas disease reactivation occurred during the study period.

Abstract

Patients with chronic Chagas disease (CD) and autoimmune rheumatic disorders (ARD) receiving immunosuppressive therapy are at risk for CD reactivation (CDR). This study monitored parasitemia over 9-121 months in six patients with CD and ARD using conventional and quantitative PCR and parasitology. Five patients showed parasitemia; two had elevated levels of parEq/mL (49-458.7). One patient received benznidazole with a marked decrease in parasitemia; in the other, treatment was discontinued after 12 days due to toxicity. No CDR occurred. These findings support the need for qPCR standardization for preemptive therapy and suggest that benznidazole may prevent CDR in patients with high parasitemia.

Full text

INTRODUCTION

Chagas disease (CD) is increasingly prevalent in urban centers across continents due to migration of chronically infected individuals to large cities in endemic and non-endemic regions1 ^,^ 2. Autoimmune diseases have been identified in 0.8-1.3% of the 27% of immigrants affected by CD in Geneva, Switzerland1 ^,^ 2.

Patients with chronic CD, typically presenting with low but persistent parasitemia3, are at risk for CD reactivation (CDR) when associated with autoimmune rheumatic disorders (ARDs), AIDS, hematologic diseases, neoplasms, or organ and tissue transplantation. CDR is a potentially life-threatening complication with high mortality3.

A refined understanding of ARD pathophysiology has led to the development of novel disease-modifying antirheumatic drugs (DMARDs), including biologics and targeted synthetic DMARDs4. These drugs interfere, to varying degrees, with T lymphocytes and macrophage functions, which are central to controlling parasite multiplication, thereby increasing parasitemia and the risk of reactivation.

First-line therapy for ARD includes glucocorticoids and conventional synthetic immunosuppressants, such as methotrexate, leflunomide, cyclosporine, azathioprine, mycophenolate mofetil, and cyclophosphamide. In patients with inadequate remission or low disease activity, advanced therapies are indicated, including targeted synthetic DMARDs, such as biologic agents (tumor necrosis factor-alpha [TNFα] inhibitors [adalimumab, certolizumab, etanercept, golimumab, and infliximab]), T-cell inhibitors (abatacept), B-lymphocyte inhibitors (rituximab), and agents with alternative mechanisms of action beyond TNFα inhibition, such as anti-CTLA4, anti-IL6, anti-IL17, anti-IL12/23, anti-CD20, and anti-Blys5-74.

A review of the literature on CD and ARD highlights several issues: (1) Ten reported cases of CDR in patients with ARD, including systemic lupus erythematosus, rheumatoid arthritis (RA), psoriatic arthritis, systemic vasculitis, and limited cutaneous sclerosis5 ^-^ 9; (2) Eight additional patients with elevated parasitemia10 ^-^ 11, suspected of having CDR; (3) Seventy-eight patients without CDR1 ^,^ 2 ^,^ 7 ^,^ 11 ^-^ 13; (4) The lack of quantitative PCR (qPCR) standardization in non-immunosuppressed and immunosuppressed patients, limiting cross-study comparisons10 ^,^ 11 ^,^ 13.

Given the potentially increased risk of CDR, this study aimed to assess qPCR as a marker for preemptive benznidazole therapy in six patients with CD and ARDs receiving biologic treatment.

CASE REPORTS

This prospective observational study included six consecutive patients with ARD and concomitant CD, followed between 2009 and 2023 at the Infectology and Rheumatology Clinics of a university hospital. Diagnosis of T. cruzi infection was confirmed using two different serological tests (ELISA, indirect immunofluorescence, hemagglutination, or chemiluminescence)14.

Parasitemia was monitored using molecular and indirect parasitological methods (blood culture, xenodiagnosis, or both), and direct microscopic examination of blood3. Conventional PCR (cPCR) amplified a 330 bp minicircle sequence using S35 and S36 primers with modifications14. qPCR was performed with TCZ3 and TCZ4 primers to amplify a 149 bp genomic fragment using SYBR Green-based qPCR14. The positivity rate of indirect parasitological methods was 19.4% in patients with chronic CD and 57.5% in HIV-coinfected patients without CDR; molecular methods showed 37.8% and 70.2% positivity, respectively, with 100% specificity14. In prior qPCR studies14, baseline levels in chronic cases reached up to 11.0 parasite Equivalents/mL (parEq/mL), whereas in coinfected patients without CDR, stable parasitemia reached up to 67.0 parEq/mL, and variable parasitemia reached up to 390 parEq/mL14.

Benznidazole (5-7 mg/kg/day for 60 days)14 was used as preemptive antiparasitic therapy in cases with increased parasitemia, defined as parEq/mL levels higher than the maximum observed in immunosuppressed patients without CDR (> 67 parEq/mL)14 by qPCR, and a relative increase compared to the patient's previous baseline control.

Inclusion criteria: (1) Confirmed diagnosis of CD by two serological tests and an autoimmune disease diagnosed by gold standard methods; (2) Evaluation of parasitemia in at least three samples using molecular methods. Exclusion criteria: Patients followed for < 6 months or those who did not undergo the recommended tests to characterize the clinical form of CD.

This study was approved by the Ethics and Research Committee of our institution (protocol numbers 1043/07 [12/05/2007] and 1298/06 [01/17/2007]). All patients provided written informed consent. Data were collected anonymously from medical records.

Six patients with ARD and CD were followed up for 9-121 months (median, 85.5 months [interquartile range 25%-75%, 64.25- 110.5]) (Table 1).

TABLE 1:Clinical profile, immunosuppressive therapy, and T. cruzi parasitemia monitoring by cPCR, qPCR, and parasitological methods in six patients with autoimmune rheumatic disorders.CaseAge SexDiagnosis, date (month/year) Clinical activityTreatment (month/year)Laboratory exams and disease activityChagas disease evaluation (clinical form)cPCR/ Blood culture/Xeno-diagnosis X month/yearqPCR/D*months follow-up/medicines BNZ P176 FRheumatic polymyalgia 01/2009. Temporal arteritis?PRE 10 mg/d (01-03 /2009) + ETA 50 mg/week (01/ 2009 to 01/2010) (Cardiac + megaesophagus). Normal ECG, Holter: 2^nd^-degree AVB, intermittent Mobitz II. 05/2009 N/B+XNU/DN

Echo-discrete diastolic dysfunction, 07/2009N/B+XN 10/2009 N/BNAXNAU/NA U/NA

MTX until 07/2010 Esophagus-hypotonia-lower sphincter: 2012 and megaesophagus: 201602.2010 N/BNXN U/DN9P2 54 FAntisynthetase-like syndrome (polymyositis 05/2010 - Raynaud phenomenon 03/2011: poly-neuropathy, pneumopathy 05/11 - Dysphagia, muscle weakness, myalgia, myopathy (biopsy) 12/2011: microangiopathyMTX - 03/2011 to 04/2012 10-25 mg/week MP - 3 g (04/2011), AZA 100 mg/d, followed by PRE 40-80 mg/d (2011), 10-60 mg (2012), suspended (03/2014)Anti-ku antibody +, 02/11 ENM chronic myopathy, peripheral polyneuropathy CPK 6895 U/L, Aldolase 83/U/L. Anti-nuclear antibody +, CPK 1841 U/L(Cardiac + megaesophagus) ECG - RBB - ASRB 2011-2017 ECG - RBB - ASRB 2011-201705/2011 +/BNAXN 07/2011+/BNXN 11/2011 +/BNXN65.4/DN 17.5/DN 2.84/DN24MPRED PRED 40 mg/d

04/2012: worsening muscle weakness RIT (07/2013) (2^nd^ cycle), PRED 20 mg/d, MTX 20 mg/dNegatives ENA, Scl70, DNA, Sm, RoHolter 2014 - AVB 2^nd^ degree, RVV +ASBB 09/201403/2012 +/BNAXNA 05/2012 +/B+XN7.55/DN 10.6/DN

RIT (01/2014), PRED suspended (03/2014)Negatives La, ACL, LAC, Jo1 (anti-histidil tRNA sintetase)Echocardiogram - LV hypertrophy - 2011-201208/2012 +/B+XN 01/2013 +/ BNXN 03/2013 +/BNX+6.2/DN 27.5/DN 40.3/DN 22*/PRED 20-25 mg/d

Failure 3 drugs, introduced RIT (07/2012) (4 cycles) - 01/2014 RIT, PRE 25 mg/d - 01/2013 Echocardiogram 2014 - akinesia (basal segment - inferior septum)09/2013 +/B+XNA 01/2014 +/B+XN1.65/DN 6.15/DN

Esophageal manometry - hypotonia, motor activity absent 09/2013 +/B+XNA 01/2014 +/B+XN1.65/DN 6.15/DN

In middle, distal segments - 201402/2015 +/BNAXNAU/DN

MTX 25 mg/week (03/2015 - 09/2016)ImprovementEsophagoscopy - Grade I10/2015 +/BNAXNA25.3/DN

Pain and lymphnodomegaly

Megaesophagus 201605/2016 +/BNAXNA76.4/DN60*/PRED 40 mg/d

• reactional lymphadenitis 10.2016-Leucocytes 2740, platelets 146000/mm^3^, 10/2016 -12 days, BNZ 300 mg/d - suspended due to fever, rash,11/2016 N/BNAXNANA/DN65* BNZ 12 days

MTX 10-30 mg/week - 2017 to 01/2019Hb 13.2 g/dL13 days leukopenia02/2017 +/BNAXNA2.19/DN

MTX 20 mg/week - 01/2018

01/2018 +/BNAXNA46.3/DN80*MTX

enlarged mediastinal, axillary lymph nodes,MTX - 10 mg/week. No PRED since 07/201605/2018: Anti-synthetase syndrome (anti-ku+) 05/2018 +/BNAXNA63.9/DN84 *MTX

interstitial pneumopathy

06/2018 +/BNAXNA7.7/DN

Pain and joint heat (04/2019), stable pneumopathy MTX increased 20 mg/day (04/2019)

04/2019 +/BNAXNA46.95/DN93 *MTX

Arthralgia and joint edema (03/2021)MTX 20 mg/day until 01/2021 MTX - PRE 03/2021

12/2019 +/BNAXNA 06.2021+/BNAXNA23.04/DN 11.12 DN 121*P357 FRheumatoid arthritis (2008) - Ac antiCCP + LEF 20 mg/d - 09/2013-10/2017 (Megaesophagus) 2013 - ECG and echocardiogram - normal01/2014 N/BNXNU/DNA

MTX 10-15 mg/week - 2011 -11/2013 2016 - Reduced peristalsis, discrete reflux, and nonspecific 03/2014 N/BNXNU/DNA

HQ 400 mg/d - 12/2013- 05/2019 dysmotility in XR of stomach, esophagus, and duodenum07/2014 N/BNXNAU/DN

MTX 03/2015 Esophagogram: increased 01/1015 N/BNAXNAU/DN

PRE 10 mg/day - 01/2016 esophageal caliber and slow02/2016 N/BNAXNAU/DNA

suspension gastric intolerance empty time05/2017 +/BNAXNAU/DN40*PRED 10 mg/d

03/2018 N/BNAXNAU/DNA

05/2019 N/BNAXNAU/DN64*P4 67 FPrimary Sjogren syndrome (2010)MTX 17.5-25 mg/week: 09/2009-02/2014, 10 mg/week until 2023Anti-nuclear antibody +(Non-cardiac form)01/2014 N/BNAXNAU/DNA

polyarthritis, rheumatoid vasculitisPRE 2.5-10 mg/day: 2009-2015, HQ 400 mg/d (02/14) MTX 10 mg/weekanti-SSA (Ro) antibody2014 - Echocardiogram early diastolic dysfunction*03/2014 N/BNXNU/DNA

interstitial pneumopathyHQ: 06/2010-02/2015anti-SSP (La) antibody2014, 2016,2017 - Normal ECGs05/2014 N/BNXN0.47/DNA3*PRED 10mg/d

neuropathyCOL 0.5 mg/d: 03/2014-02/2015Rheumatoid factor - Negative2016 - No megaesophagus 10/2014 N/BNXNU/DN

xerostomiaPRE was reintroduced in 04/2016 CCP - Negative 01/2016 N/BNXNU/DNA

xerophthalmiapneumopathy

09/2016 N/BNXNU/DN

xerotrachea

05/2017 N/BNXNNA/DN

07/2018 N/BNXNU/DN

03/2019 N/BNXNNA/DN

05/14-2023: HQ 400 mg +

10/2020 N/BNXNNA/DN

PRE 7.5 mg/d

11/2022 N/BNXNU/DNA106*P5 62 FRheumatoid arthritis (2005)CSP 100 mg/d (10/2009-06/2013). MTX 25 mg/week (2005-2014) Rheumatoid factor + DAS 28: 7.88 - 06/2013 (Cardiac form)01/2014 N /BNXNAU/DNA

Rheumatoid vasculitisLEF-neuropathy (2009) ABA (12/2013-03/2014)SDAI 5, CDAI 5, AVG 50 DAS 28: 6.1 - 12/20132015, 2019 - Frequent atrial extrasystole ECG. 10/2014 N /BNAXNA U/DN

Treated pulmonary tuberculosisPRE 5-20 mg/day (10/2009 - 11/2014) Echocardiogram discrete alteration of left ventricular10/2015 N /BNAXNAU/DN

Mild obstructive pulmonary disease, diabetes mellitus (II)TOC (08/2014-08/2018). (10/2014) - TOC, ME 25 mg/week, PRE 5 mg/day relaxation. 2020 - Holter- non-sustained atrial tachycardia and rare 05/2018 N /BNAXNA NA/DNA

Heart failure (07/2019)TOC (08/2014-08/2018). (2019) - TOC 4/4 weeks, PRE 10 mg/d, MTX 10 mg/dDAS 28: 6.6 - 08/2014episodes of aberrant intraventricular conduction.06/2019 N/BNAXNA NA/DNA65*P6 M43 FRheumatoid arthritis (2000) MTX, HQ, RIT before 2010 -2011, TOC (2011)Positive rheumatoid factor C-reactive protein: 46 mg/L(Cardiac form) 2014- RBB, junctional escape, low voltage, changeable 04/2014 +/BNAXNA409,5/DNAZA, PRE 10 mg/d

Desquamative pneumonitisPRE 10 mg/d (2012) - 15 mg/day (10/2017) DAS 28: 6.05 (10/2012) Atrial pacemaker - ECG09/2014 +/B+XNA29.5/DN

12/2020 - Pulmonary biopsyLEF 2012-06/2013 (neuropathy)DAS 28: 4.79 (07/2014) 2014 - Rare atrial and ventricular 10/2014 +/B+XNA6.3/DN

Chronic non-specifc interstitial pneumoniaAZA 100-150 mg/d (12/2012 -09/2016)DAS 28: 5.43 (04/2015); 7.05 (06/2015); 3.8 (10/2015)Extrasystoles, and supraventricular tachycardias 03/2015 +/BNAXNA51.0/DN*11/AZA, ABA, PRE 10 mg/d

Progressive worsening of interstitial pneumoniaABA 500 mg (09/2014-10/ 2015)DAS 28: 6.4 (11/2015)2014 - Left ventricular ejection fraction 60% - Echocardiogram Esophagoscopy - Normal10/2015 +/BNAXNA223.0/DN18*/AZA,ABA, PRE 10 mg/d

CT scans and digital clubbing, TCO (05/2015-02/2016) - FalhaDAS 28: 6.4 (02/2016) 2013-2017 - Chest Xray - interstitial pneumopathy associated with ARD02/2016 +/BNAXNA458.7/DN22*/AZA.TOC, PRE 10 mg/d

interstitial pneumopathyTOF (10/2015-02/2016)

04/2016 N/BNAXNAU/DN24*/BNZ 03.2016 30d

AZA, PRE (09/2016)

07/2016 +/BNAXNAU/DN

RIT (11/2017)

09/2016 N/BNAXNAU/DNA

PRE 15 mg (02/2018)DAS: 3.8 (02/2018) 05/2017 +BNAXNAU/DN

MP 100 mg (02/2018)PCR 46 01/2018 +/BNAXNAU/DN45*/MP

RIT (11/2018 and 08/2022-09/2023)

06/2018 +/BNAXNA0.08/DN

TOC (03/2019-05/2022)

12/2018 +/BNAXNA0.40/DN

TOC (03/2020-02/2022) hepatotoxicity 03/2019 +/BNAXNAU/DN

PRE 10-20 mg/d (08/2020-05/2022) - ?40 mg /day+MPREDSevere evolution pneumopathy, arthralgia, 12/2020 +/BNAXNA0.12/DN80*MP+ PRE 40 mg/d

PRED 40 mg/d (08/2022) dyspnea lung transplant no 09/2022 +/BNAXNA3.1/DN

Lung amyloidosisMP 100 mg (08/2022) dyspnea: MMF (12/2022) toxicityIndicated: renal insufficiency 12/2022 +/BNAXNAU/DN104*MP

Renal amyloidosisPRE low doses (09/2023)

08/2023 +/BNAXNA2.3/DN112* F: female; M: male; (y): years; cPCR: conventional polymerase chain reaction (qualitative); qPCR: quantitative PCR; U: undetectable; D: direct parasitology; *months of follow-up; PRE: prednisone; MP: methylprednisolone; MTX: methotrexate; ETA: etanercept; AZA: azathioprine; CSP: cyclosporine; COL: colchicine; RIT: rituximab; HQ: hydroxychloroquine; LEF: leflunomide; ABA: abatacept; TOC: tocilizumab; TOF: tofacitinib; N: negative; ECG: electrocardiogram; Echo: echocardiogram; B: blood culture; NA: not available; X: xenodiagnosis; D: direct microscopy; BNZ: benznidazole; ku: Ku70/Ku80; DNA: binding protein; CPK: creatine phosphokinase; CRP: C-reactive protein; RBB: right branch block; ASBB: anterosuperior branch block; AVB: atrioventricular block; CCP: cytric citrullinated peptide; CPK: creatine phosphokinase; anti ENA: nuclear extrated antibodies (RNP, Sm, SSa/Ro, SSb/L); anti Ro (SSa) and La (SSb): anti RNA antibodies; Scl70-anti-toporoisomerase 1, antiI-SSm: anti-Smith antibody; aCL: anticardiolipin; LAC: lupus anticoagulant; anti JO: anti-histidine tRNA synthetase; SDAI: simple disease activity index; CDAI: clinical disease activity index; DAS: disease activity score.

All patients were female, aged 43-76 years; two had cardiopathy, two had cardiopathy with megaesophagus, one had megaesophagus, and one had a non-cardiac form of CD^3^ (Table 1).

Five patients had T. cruzi parasitemia confirmed by parasitological or molecular method14, and two showed elevated parasitemia by qPCR (76.4 and 458.7 parEq/mL). The first patient (P2) with severe polymyositis and antisynthetase syndrome received methylprednisolone followed by prednisone (>20 mg/day) during 2/5 relapses, combined with methotrexate, or rituximab and methotrexate alone in three relapses. The second patient (P6) with RA received prednisone (10-15 mg/day) and azathioprine (100 mg/day) in four relapses, combined with azathioprine in one relapse, abatacept in two, and tocilizumab in one.

Patient 1

A 76-year-old female patient was diagnosed with rheumatic polymyalgia and temporal arteritis and treated with prednisone, methotrexate, and etanercept without clinical improvement. CD progressed to cardiopathy (2011) and megaesophagus (2016). Parasitemia remained negative by direct microscopy, cPCR, qPCR, blood culture, and xenodiagnosis for 9 months, except for two positive blood cultures in 2009.

Patient 2

A 54-year-old female was diagnosed with polymyositis and antisynthetase syndrome associated with pneumopathy. She was prescribed methotrexate, methylprednisolone, azathioprine, prednisone (up to 60 mg/day), cyclosporine, and rituximab.

CD had progressed to cardiopathy and megaesophagus*. T. cruzi* parasitemia remained positive for 10 years.

Antiparasitic treatment was considered due to increasing parasitemia despite severe myopathy. With an increase to 76.4 parEq/mL from basal levels of 1.65-7.7 parEq/mL, benznidazole (5 mg/kg/day) was prescribed on October 2016 but discontinued after 12 days due to fever and generalized erythema; leukopenia was observed the day after discontinuation. Parasitemia was monitored thereafter, with transient elevations up to 63.9 parEq/mL in the absence of an effective alternative antiparasitic drug with no toxic effects.

Patient 3

A 57-year-old female was diagnosed with RA in 2008 and treated with prednisone, leflunomide, methotrexate, and hydroxychloroquine. She had Chagasic megaesophagus. T. cruzi parasitemia was monitored for 64 months, with one of eight cPCR results testing positive.

Patient 4

A 67-year-old female was diagnosed with Sjögren’s syndrome, polyarthritis, rheumatoid vasculitis, interstitial pneumopathy, and bilateral keratitis. She was prescribed methotrexate, prednisone, and hydroxychloroquine. CD was classified as non-cardiac. T. cruzi parasitemia was monitored for 106 months, with only 1/7 qPCRs showing a low par Eq/mL level (Table 1).

Patient 5

A 62-year-old female with RA (2005) was treated with methotrexate, cyclosporine, abatacept, prednisone, tocilizumab, and hydroxychloroquine. A cardiac form of CD was diagnosed, and parasitemia remained negative for 65 months.

Patient 6

A 43-year-old female was diagnosed with RA associated with desquamative pneumonitis. She had previously received methotrexate and hydroxychloroquine, followed by rituximab, and was later switched to tocilizumab. The patient was subsequently prescribed leflunomide, azathioprine, abatacept, and tofacitinib.

Cardiac CD was confirmed. Parasitemia monitoring revealed transient peaks between April 2014 and October 2015. In February 2016, 458.7 parEq/mL was detected, and benznidazole (300 mg/day) was prescribed for 60 days. Parasitemia decreased to undetectable or very low qPCR levels, with no recurrence of high parasitemia, despite high-dose methylprednisolone use during the 84-month follow-up post-therapy.

DISCUSSION

This study found positive parasitemia in 83.3% of patients (5/6), which was higher than that observed in chronic non-immunosuppressed cases14 and comparable to that in T. cruzi-HIV coinfected patients14. This frequency also exceeded the 53.8% reported in one case series11, while such data were not available in other series1 ^,^ 2 ^,^ 7 ^,^ 11.

No cases of CDR were identified according to standard parasitological or quantitative molecular methods, contrasting with the 11.8% (6/51) reported previously7 (Table 1). In most cases, corticosteroids > 20 mg/day were not prescribed for extended periods as first-line therapy, as they interfere with cellular and humoral immune response4.

In two patients, parasitemia levels of 65.4 parEq/mL (P2) and 409.5 parEq/mL (P6) at the beginning of follow-up were transient and decreased to 2.84 and 6.3 parEq/mL, respectively, without antiparasitic treatment (Table 1). No clinical signs, symptoms, or positive direct parasitological methods indicative of CDR were observed.

In ARD, comparing high parasitemia levels across studies is challenging due to the lack of qPCR standardization. Parasitemia levels of 57.413 and > 50 parEq/mL^11^ have been reported as markers of CDR, whereas levels around 1,000 copies were considered low and not indicative of CDR10. In contrast, high values ranging from 996 to 5,575 parEq/mL^11^ may suggest CDR.

Standardization of qPCR as a marker for CDR has been infrequently reported across different laboratories in patients with CD with varying levels of immunosuppression (chronic CD and immunosuppressed CD patients with or without CDR)14. In patients with chronic CD, the maximum parasitemia detected was 11.0 mEq/mL. Among CD-HIV coinfected patients without CDR, parasitemia reached up to 67.0 mEq/mL in those with stable levels, and 390 parEq/mL in a group with variable parasitemia14.

Notably, preemptive antiparasitic therapy in P6 was followed by a marked decrease in parasitemia to undetectable or 2.3 parEq/mL during the long follow-up period, as reported in patients with high parasitemia (other ARD patients with CD11, T. cruzi HIV coinfection14, stem cell transplantation, and medullary aplasia without CDR15 ^-^ 16). None of these cases developed CDR. However, the toxicity of antiparasitic drugs must be considered, particularly in patients with systemic myopathy (P2). No similar long follow-up data after treatment have been reported.

In this context, based on prior studies13 ^-^ 16 and the present series, absolute qPCR levels ≥ 70 -100 parEq/mL or increased parasite levels relative to a patient's baseline may indicate the need for preemptive therapy, warranting confirmation in larger cohorts of patients with of ARD and CD.

CONCLUSION

This study confirmed the utility of qPCR as a marker of increased parasitemia. A marked and sustained reduction in qPCR levels was observed following benznidazole therapy, despite subsequent high-dose corticosteroid use. Unlike previous reports, no cases of CDR occurred, possibly due to the less frequent use of high-dose corticosteroids for long periods.