article: There is a paucity of published reports on pregnancy outcome following scrub and murine typhus despite these infections being leading causes of undifferentiated fever in Asia. This study aimed to relate pregnancy outcome with treatment of typhus. Data were analyzed from: i) pregnant women with a diagnosis of scrub and/or murine typhus from a fever cohort studies; ii) case series of published studies in PubMed using the search terms “scrub typhus” (ST), “murine typhus” (MT), “Orientia tsutsugamushi”, “Rickettsia tsutsugamushi”, “Rickettsia typhi”, “rickettsiae”, “typhus”, or “rickettsiosis”; and “pregnancy”, until February 2014 and iii) an unpublished case series. Fever clearance time (FCT) and pregnancy outcome (miscarriage and delivery) were compared to treatment. Poor neonatal outcome was a composite measure for pregnancies sustained to 28 weeks or more of gestation ending in stillbirth, preterm birth, or delivery of a growth restricted or low birth weight newborn. There were 26 women in the fever cohort. MT and ST were clinically indistinguishable apart from two ST patients with eschars. FCTs (median [range] hours) were 25 [16–42] for azithromycin (n = 5), 34 [20–53] for antimalarials (n = 5) and 92 [6–260] for other antibiotics/supportive therapy (n = 16). There were 36. 4% (8/22) with a poor neonatal outcome. In 18 years, 97 pregnancies were collated, 82 with known outcomes, including two maternal deaths. Proportions of miscarriage 17. 3% (14/81) and poor neonatal outcomes 41. 8% (28/67) were high, increasing with longer FCTs (p = 0. 050, linear trend). Use of azithromycin was not significantly associated with improved neonatal outcomes (p = 0. 610) The published ST and MT world literature amounts to less than 100 pregnancies due to under recognition and under diagnosis. Evidence supporting the most commonly used treatment, azithromycin, is weak. Collaborative, prospective clinical trials in pregnant women are urgently required to reduce the burden of adverse maternal and newborn outcomes and to determine the safety and efficacy of antimicrobial treatment.
  The true burden of scrub typhus (ST) and murine typhus (MT) in South-east Asia remains largely unknown as diagnostic tests are rarely used or available [1]. Still less is known about typhus in pregnancy with no (large scale) epidemiological studies addressing the subject in this group [2]. No data suggests that typhus occurs more or less commonly in pregnancy. Studies showing that the malaria vector An. gambiae mosquitoe was more attracted to pregnant women than non-pregnant women [3], have not been performed with the vectors of ST (mites) and MT (fleas). There is a scatter of small case series and reports on ST and MT [4]–[16] which tend to suggest serious repercussions for the mother and fetus, and congenital transmission has also been reported [10], [11]. There is a definitive lack of new studies or data in the published literature with a four year gap between the two latest publications [16], [17]. This a likely reflection of the difficulty of confirming the diagnosis and following a woman treated in out- or in-patients through to pregnancy outcome in the obstetric department. The tetracycline-class of antibiotics, principally doxycycline, treat rickettsial diseases [18]. Congenital malformation has not been associated with doxycycline use [19], [20] but the class effect ascribed to tetracyclines including possible effects on the musculoskeletal development of the fetus [21] and staining of teeth in young children [22] precludes its use. While short courses of doxycycline are probably safe there is insufficient evidence to support their use as first line therapy [5], [23]. Macrolide antibiotics such as azithromycin, are safe in pregnancy and reported to be equally effective to doxycycline for the treatment of scrub typhus [24]. The aim of this study was to undertake a detailed analysis of ST and MT cases from a previously published fever cohort and of the published literature to provide further detail on pregnancy outcomes in relation to treatment.
  For women participating in the fever cohort the patient information sheet and consent form were available in Karen and Burmese languages. Willing participants signed (literate) or provided a thumb-print (illiterate) on the consent form. Approval for the study was granted by the Oxford Tropical Research Ethics Committee, UK (#013-03) [16]. Details of the methods can be found in this previously published study on the epidemiology of fever in refugee and migrant pregnant women on the Thai-Myanmar border where scrub and murine typhus accounted for 12. 3% (26/211) of fever diagnoses in 203 women [16]. Briefly, febrile (aural temperature >37. 5°C) pregnant women following the antenatal clinics of Shoklo Malaria Research Unit (SMRU) were offered a fever screen. Relevant to this study is the diagnosis typhus and malaria. Typhus was confirmed by PCR [25]–[28] and/or in vitro isolation of Rickettsia spp. [29] and/or positive reference serology measured by four-fold rise in paired sera IFA [30], [31] to define acute infection as MT (Rickettsia typhi) or ST (Orientia tsutsugamushi). Malaria was confirmed by microscopy of thick and thin malaria smears, stained with Giemsa and examined under oil immersion. Smears were only declared negative after 200 fields were read. None of the ST and MT results were available for the patient as these were initially stored and later analyzed off-site. Women were admitted and treatment was initiated according to the clinical examination and available laboratory test results. This implies patients may have been treated for other infections such as malaria because the result was immediately available and consistent with the clinical picture. ST or MT may not have treated as it was not suspected and the laboratory result was unavailable. Patients were monitored for fever clearance time (FCT) by 6-hourly aural (Genius) temperature measurement. Women followed regular antenatal care after discharge and were encouraged to deliver with skilled birth attendants where they are weighed directly after birth. Infants born at home and not weighed on the day of birth had their weight adjusted as described for this population [32]. Poor neonatal outcome was a composite measure for women whose pregnancy was sustained to 28 weeks or more of gestation and included stillbirth, prematurity and fetal growth restriction defined as a birth weight for gestational age of <10th centile for the population [33]. Since Dec 2012 pregnant women at SMRU with fever have routinely been tested with the SD Bioline Tsutsugamushi from Standard Diagnostics, Korea, a rapid test, in addition to other standard fever screening tests. This rapid diagnostic test is non-specific testing IgG, IgM and IgA Orientia tsutsugamushi antibodies. Acute and convalescent serum was collected. Remaining serum was stored at −20°C. These samples were collected over a one year period and processed in the same laboratory as per the aforementioned fever cohort women. ST was confirmed by PCR [25]–[28] and/or positive reference serology measured by four-fold rise in paired sera IFA [30], [31]. Women were followed up in the routine antenatal service and encouraged to birth with skilled birth attendants at Shoklo Malaria Research Unit. Of 25 women with a positive rapid test there were three with a confirmed ST diagnosis and a known pregnancy outcome included for analysis. The pooling of data into a single data set includes the aforementioned fever cohort women, additional unpublished cases, and data reported from case reports or case series in 16 different publications [5]–[7], [9], [10], [12], [15]–[17], [34] (Supporting Information S1). As the population birth weight percentiles were unknown for these publications, low birth weight (<2500 g) replaced fetal growth restriction in the definition of poor neonatal outcome. Individual patient data including age, gestation at infection, days of fever on admission, FCT, treatment, maternal death, birth outcome (including miscarriage (delivery before viability set at <28 weeks for resource limited settings), delivery and unknown), birth weight and gestational age at delivery, live birth or stillbirth, birth weight and congenital normality were extracted from published studies into a data spreadsheet (Supporting Information S2). Data were described using the statistical program SPSS Statistics version 20. 0 for Windows (SPSS Benelux inc. , Gorinchem, Netherlands). Continuous normally distributed data were described by the mean (standard deviation, range) and non-normally distributed data by the median [min-max] and comparisons used the Student' s t-test or Mann-Whitney U test, respectively. The number and percentage were given for categorical data and compared using the X2-test with Yates' correction, or the Fisher' s exact test when applicable. Poor neonatal outcome was compared by univariate analysis to the treatment group. Given the small sample size and to avoid erroneous conclusions no modeling of poor neonatal outcome was attempted.
  From 2004 to 2006,26 febrile pregnant women were identified as having evidence of rickettsial infection with confirmation by serology in 9 women, serology and PCR in 17 women (MT n = 7, ST n = 9, and both n = 1). In 4 of these cases O. tsutsugamushi was isolated from blood culture and all of these women were PCR and serology positive. Culture positive cases included the single patient with mixed ST and MT infection. Note that in 5 cases there was concomitant infection with malaria. Malaria cases were all microscopic: three MT (with two concomitant P. falciparum and one P. vivax infections) and two ST with concomitant P. falciparum. These five patients were grouped respectively into MT and ST [16]. The patient who had dual infection with ST and MT (dynamic serology for MT, and PCR positivity for both) documented at 37+2 weeks+days gestation could not be placed into either group (ST or MT) for analysis. Briefly she was a 33 year old with 4 previous births, a FCT of 119 hours with IV ceftriaxone and a normal birth outcome (singleton, estimated gestation 41+3 weeks+days and birth weight of 3600 g). Overall there were 13 women with ST, 12 with MT and one with ST and MT. The baseline demographic and obstetric characteristics have been detailed for MT and ST diagnostic groups (Table 1). There were no significant differences observed in baseline characteristics. Of note is the higher proportion (non-significant): of flea-borne MT was higher in the refugee camp than in migrants; and of smokers in the ST group. From Dec 2012 until Feb 2014 there were 22 febrile pregnant women with a positive SD Bioline Tsutsugamushi RDT and serum samples available for further diagnostic testing. Of these women only three had a 4-fold rising ST IgM titre (two 1∶400 to 1∶1600 and one 1∶1600 to 1∶6400) and none had a positive ST PCR result. All 3 women delivered term, live born, normal infants one of which was of low birth weight (treated with paracetamol and fever had cleared by the time the lab result was available). Treatment in the other two women was azithromycin in one case based on protocol, and multiple antibiotics for the other case: initially ciprofloxacin for suspected pyelonephritis, two days later erythromycin for suspected exacerbation of asthma and one day later azithromycin based on the positive RDT result with rapid fever clearance, <24 hours, once this drug was commenced. Over an 18 year period, from 1992 to 2014 there were 97 women with a diagnosis of MT or ST (or both) in pregnancy including the 26 reported from the SMRU fever cohort [16] described above, the 3 unpublished cases, and 85 in peer reviewed publications [5]–[7], [9], [10], [12], [15]–[17], [34] (Supporting Information S1). The number of cases (per publication) included 42 (n = 1), 26 (n = 1), 9 (n = 1), 5 (n = 2), 2 (n = 2) and 1 (n = 3), from five different countries including 52 from India, 30 from the Thai-Myanmar border, 11 from Korea, and 1 from Taiwan. The three unpublished cases were also from the Thai-Myanmar border. Reported diagnostic methods were predominantly serology-based, which included Weil-Felix and IFA testing, and few were paired titrations and/or PCR (Supporting Information S1).
  The main limitations of this study are the use of retrospective data subject to reporting bias and the weak serology-based rickettsial diagnostics of the case series [54]. Despite this the proportion of poor neonatal outcome was similar in the fever cohorts with more reliable confirmation including PCR (Table 5). Only real-time diagnostic capacity, classification of disease severity and standardized approaches to treatment in prospectively followed pregnancies will allow clarification of the findings [55]. Given that typhus represents the leading cause of treatable undifferentiated fever in Southeast Asia [56], [57], and that its regions of high endemicity are among the most populous areas in the world, the need for prospective fever-in-pregnancy studies to estimate the burden and best treatment of disease in this group is obvious. Asymptomatic infection is also a high possibility and the implications for pregnancy unknown. Theoretically azithromycin is the drug of choice in pregnancy but the worldwide data collated here only provides support for level-4 evidence for the use of this drug and this must improve. The data presented here highlights the potential implications and severity of this easily treatable infection and demonstrates the inexorably slow pace of improvement in our understanding of rickettsial illness in pregnancy: less than 100 women with a known pregnancy outcome in 18 years. Progress towards the 2015 countdown to Millenium Development Goals to improve maternal, newborn and child survival will not be met in relation to scrub and murine typhus. Evidence to support the use of azithromycin is weak. The correct antimicrobial or combination for undifferentiated fever in pregnant women in South-east Asia is unknown.
summary: Typhus is an under-recognised and under-studied public health problem in Asia. In rural areas of Southeast Asia murine and scrub typhus are probably the most common treatable cause of fever. The estimated number of scrub typhus cases in Southeast Asia, more than 1 million yearly, results in approximately 50–80,000 deaths per year. Treatment delays due to lack of appropriate diagnostics and lack of awareness lead to a substantial health and economic impact in the one of the world' s most densely populated regions. Only 97 cases in pregnancy are available from the published world literature over the past 18 years. Only 82 of these had known outcomes, including two maternal deaths. The proportion of poor neonatal outcome including stillbirth, prematurity and low birth weight was high occurring in more than 40% of pregnancies, and higher when the fever clearance time was longer. While poor neonatal outcomes were observed with all antibiotics prescribed, azithromycin appeared to be associated with shorter fever clearance times but this was not statistically significant. Evidence to support the use of azithromycin is weak. The correct antimicrobial or combination for undifferentiated fever in pregnant women in Southeast Asia is unknown.
keywords: relapsing fever
  neonatology
  bacterial diseases
  infectious diseases
  miscarriage and stillbirth
  veterinary diseases
  zoonoses
  medicine and health sciences
  women's health
  maternal health
  pregnancy
  neglected tropical diseases
  typhus of rickettsiae
  biology and life sciences
  tropical diseases
  veterinary science
  pediatrics
title: Pregnancy Outcome in Relation to Treatment of Murine Typhus and Scrub Typhus Infection: A Fever Cohort and a Case Series Analysis