Predominance of multidrug-resistant Salmonella Typhi genotype 4.3.1 with low-level ciprofloxacin resistance in Zanzibar.

BACKGROUND: Typhoid fever is a common cause of febrile illness in low- and middle-income countries. While multidrug-resistant (MDR) Salmonella Typhi (S. Typhi) has spread globally, fluoroquinolone resistance has mainly affected Asia. METHODS: Consecutively, 1038 blood cultures were obtained from patients of all age groups with fever and/or suspicion of serious systemic infection admitted at Mnazi Mmoja Hospital, Zanzibar in 2015-2016. S. Typhi were analyzed with antimicrobial susceptibility testing and with short read (61 strains) and long read (9 strains) whole genome sequencing, including three S. Typhi strains isolated in a pilot study 2012-2013. RESULTS: Sixty-three S. Typhi isolates (98%) were MDR carrying blaTEM-1B, sul1 and sul2, dfrA7 and catA1 genes. Low-level ciprofloxacin resistance was detected in 69% (43/62), with a single gyrase mutation gyrA-D87G in 41 strains, and a single gyrA-S83F mutation in the non-MDR strain. All isolates were susceptible to ceftriaxone and azithromycin. All MDR isolates belonged to genotype 4.3.1 lineage I (4.3.1.1), with the antimicrobial resistance determinants located on a composite transposon integrated into the chromosome. Phylogenetically, the MDR subgroup with ciprofloxacin resistance clusters together with two external isolates. CONCLUSIONS: We report a high rate of MDR and low-level ciprofloxacin resistant S. Typhi circulating in Zanzibar, belonging to genotype 4.3.1.1, which is widespread in Southeast Asia and African countries and associated with low-level ciprofloxacin resistance. Few therapeutic options are available for treatment of typhoid fever in the study setting. Surveillance of the prevalence, spread and antimicrobial susceptibility of S. Typhi can guide treatment and control efforts.

Early to mid-pregnancy HbA1c levels and its association with adverse pregnancy outcomes in three low middle-income countries in Asia and Sub-Saharan Africa.

BACKGROUND: Hyperglycemia during pregnancy leads to adverse maternal and fetal outcomes. Thus, strict monitoring of blood glucose levels is warranted. This study aims to determine the association of early to mid-pregnancy HbA1c levels with the development of pregnancy complications in women from three countries in South Asia and Sub-Saharan Africa. METHODS: We performed a secondary analysis of the AMANHI (Alliance for Maternal and Newborn Health Improvement) cohort, which enrolled 10,001 pregnant women between May 2014 and June 2018 across Sylhet-Bangladesh, Karachi-Pakistan, and Pemba Island-Tanzania. HbA1c assays were performed at enrollment (8 to < 20 gestational weeks), and epidemiological data were collected during 2-3 monthly household visits. The women were followed-up till the postpartum period to determine the pregnancy outcomes. Multivariable logistic regression models assessed the association between elevated HbA1c levels and adverse events while controlling for potential confounders. RESULTS: A total of 9,510 pregnant women were included in the analysis. The mean HbA1c level at enrollment was found to be the highest in Bangladesh (5.31 ± 0.37), followed by Tanzania (5.22 ± 0.49) and then Pakistan (5.07 ± 0.58). We report 339 stillbirths and 9,039 live births. Among the live births were 892 preterm births, 892 deliveries via cesarean section, and 532 LGA babies. In the multivariate pooled analysis, maternal HbA1c levels of ≥ 6.5 were associated with increased risks of stillbirths (aRR = 6.3, 95% CI = 3.4,11.6); preterm births (aRR = 3.5, 95% CI = 1.8-6.7); and Large for Gestational Age (aRR = 5.5, 95% CI = 2.9-10.6). CONCLUSION: Maternal HbA1c level is an independent risk factor for predicting adverse pregnancy outcomes such as stillbirth, preterm birth, and LGA among women in South Asia and Sub-Saharan Africa. These groups may benefit from early interventional strategies.

Proteomic analysis of peripheral blood mononuclear cells isolated from patients with pulmonary tuberculosis: A pilot study from Zanzibar, Tanzania.

This study aimed at exploring the proteomic profile of PBMCs to predict treatment response in pulmonary tuberculosis (PTB). This was a pilot study conducted among 8 adult patients from Zanzibar, Tanzania with confirmed PTB. Blood samples were collected at baseline, at 2 months of treatment, and at the end of treatment at 6 months. Proteins were extracted from PBMCs and analyzed using LC-MS/MS based label free quantitative proteomics. Overall, 3,530 proteins were quantified across the samples, and 12 differentially expressed proteins were identified at both 2 months of treatment and at treatment completion, which were involved in cellular and metabolic processes, as well as binding and catalytic activity. Seven were downregulated proteins (HSPA1B/HSPA1A, HSPH1, HSP90AA1, lipopolysaccharide-binding protein, complement component 9, calcyclin-binding protein, and protein transport protein Sec31A), and 5 proteins were upregulated (SEC14 domain and spectrin repeat-containing protein 1, leucine-rich repeat-containing 8 VRAC subunit D, homogentisate 1,2-dioxygenase, NEDD8-activating enzyme E1 regulatory subunit, and N-acetylserotonin O-methyltransferase-like protein). The results showed that proteome analysis of PBMCs can be used as a novel technique to identify protein abundance change with anti-tuberculosis treatment. The novel proteins elucidated in this work may provide new insights for understanding PTB pathogenesis, treatment, and prognosis.

Malaria prevalence and performance of diagnostic tests among patients hospitalized with acute undifferentiated fever in Zanzibar.

BACKGROUND: Control efforts in Zanzibar reduced the burden of malaria substantially from 2000 to 2015, but re-emergence of falciparum malaria has been observed lately. This study evaluated the prevalence of malaria and performance of routine diagnostic tests among hospitalized fever patients in a 1.5 years period in 2015 and 2016. METHODS: From March 2015 to October 2016, paediatric and adult patients hospitalized with acute undifferentiated fever at Mnazi Mmoja Hospital, Zanzibar were included. The malaria prevalence, and performance of rapid diagnostic test (RDT) and microscopy, were assessed using polymerase chain reaction (PCR) as gold standard. RESULTS: The malaria prevalence was 9% (63/731). Children under 5 years old had lower malaria prevalence (5%, 14/260) than older children (15%, 20/131, p = 0.001) and persons aged 16 to 30 years (13%, 15/119, p = 0.02), but not different from persons over 30 years old (6%, 14/217, p = 0.7). All cases had Plasmodium falciparum infection, except for one case of Plasmodium ovale. Ten malaria patients had no history of visiting mainland Tanzania. The RDT had a sensitivity of 64% (36/56) and a specificity of 98% (561/575), and microscopy had a sensitivity of 50% (18/36) and  a specificity of 99% (251/254), compared to PCR. The malaria parasitaemia was lower in patients with false negative results on RDT (median 7 × 103 copies/µL, interquartile range [IQR] 2 × 103 - 8 × 104, p = 0.002) and microscopy (median 9 × 103 copies/µL, IQR 8 × 102 - 7 × 104, p = 0.006) compared to those with true positive RDT (median 2 × 105 copies/µL, IQR 3 × 104 - 5 × 105) and microscopy (median 2 × 105 copies/µL, IQR 6 × 104 - 5 × 105). CONCLUSIONS: The study emphasizes that malaria was a frequent cause of febrile illness in hospitalized patients in Zanzibar in the years 2015-2016, particularly among school age children and young adults. We found evidence of autochthonous malaria transmission in Zanzibar. Compared to PCR, both RDT and microscopy had low sensitivity, and false negative results were associated with low parasitaemia. While low parasitaemia identified only by PCR in a semi-immune individual could be coincidental and without clinical relevance, clinicians should be aware of the risk of false negative results on routine tests.

Evaluation of treatment response in extrapulmonary tuberculosis in a low-resource setting.

BACKGROUND: Diagnosing extrapulmonary tuberculosis (EPTB) is challenging and many patients are initiated on empirical anti-TB treatment without a laboratory confirmed diagnosis. Monitoring treatment response is thus important to ensure correct diagnosis and proper disease management. The definition of satisfactory response to treatment in EPTB remains unclear. The objectives of this study were to describe the clinical presentation of EPTB and the effect of treatment on clinical parameters. Further, to assess if simple clinical parameters, without laboratory data, could evaluate treatment response. METHODS: Prospective cohort study of presumptive EPTB patients at Mnazi Mmoja Hospital, Zanzibar. By using a composite reference standard, patients were categorized as TB or non-TB cases. The TB patients were followed during anti-TB treatment. RESULTS: There were 64 TB and 62 non-TB cases. The frequency of symptoms at baseline were comparable in TB and non-TB patients, with lymphadenitis and pleuritis as the most common manifestations. Among TB cases, there was a trend towards regression of lymphadenopathy after 2 months, and at treatment completion 24/28 (86%) cases showed full regression. Weight gain ≥5% was reported in 36/49 (73%) of the TB patients at 2 months and in 38/46 (83%) at treatment completion. After 2 months of treatment, a combination of clinical parameters; improvement of symptoms (50/50), ≥5% weight gain (36/49) and regression of physical signs (45/49) correlated with the treatment response. CONCLUSIONS: An algorithm including only simple clinical parameters could be used as an easy tool to assess treatment responses in low-resource settings. However, this needs to be tested on a larger sample size.

IP-10 dried blood spots assay monitoring treatment efficacy in extrapulmonary tuberculosis in a low-resource setting.

Treatment efficacy is difficult to evaluate in extrapulmonary tuberculosis (EPTB) patients. Interferon-γ inducible protein (IP-)10 has been suggested as a biomarker for response to treatment. We have investigated if IP-10 from dried plasma spots (DPS) or dried blood spots (DBS) can be used in treatment monitoring of EPTB patients in a low-resource setting of Zanzibar. IP-10 levels in plasma, DPS and DBS samples collected before, during (2 months) and after TB treatment of 36 EPTB patients (6 culture and/or Xpert MTB/RIF positive and 30 clinically diagnosed) and 8 pulmonary tuberculosis (PTB) patients, were quantified by an enzyme-linked immunosorbent assay. There was a high positive correlation between IP-10 measured in plasma and DPS and DBS, respectively. We found a significant decline in IP-10 levels from baseline to end of treatment in plasma, DPS and DBS, both in EPTB and PTB patients. The declines were observed already after 2 months in HIV negative patients. In conclusion, the DPS/DBS IP-10 assay allows for easy and manageable monitoring in low-resource settings and our findings suggest that IP-10 may serve as a biomarker for treatment efficacy in EPTB patients, albeit further studies in cohorts of patients with treatment failure and relapse are needed.

Diagnostic delay in extrapulmonary tuberculosis and impact on patient morbidity: A study from Zanzibar.

BACKGROUND: Early and proper treatment of tuberculosis could have an important impact on the morbidity, mortality and the economic situation of patients. There is insufficient knowledge on the extent of diagnostic delay and the associated factors in extrapulmonary tuberculosis (EPTB). The aims of this study were to assess the health care seeking behaviour, EPTB knowledge and diagnostic delay in presumptive EPTB patients at the main referral hospital in Zanzibar, factors associated with longer delay, and the impact of untreated EPTB on self-rated health. MATERIALS AND METHODS: Prospective data collection using a semi-structured questionnaire in patients presenting with symptoms suggestive of EPTB. The time between the onset of symptoms and first visit to a health care provider (patient delay), and then to the initiation of treatment (health system delay) and total delay were analysed according to sociodemographic and clinical factors and health care seeking trajectories. The EQ-5D-3L was used among the adult EPTB patients to assess the impact of treatment on self-rated health. RESULTS: Of the 132 patients with median age of 27 years (interquartile range 8-41), 69 were categorized as TB cases and 63 as non-TB cases. The median patient, health system and total delays were 14, 34 and 62 days respectively, among the EPTB patients. A longer health system delay with repeated visits to the same health care level was reported. Significantly better self-rated health status was described after treatment. The knowledge regarding extrapulmonary disease was low. CONCLUSION: Many EPTB patients, presenting to the main referral hospital in Zanzibar, experience a long delay in the initiation of treatment, specially patients with TB lymphadenitis. The health system delay is the major contributor to the total delay. The improvement of self-rated health after treatment implies that timely treatment has the potential to reduce morbidity and the economic loss for the patient.

MPT64 antigen detection test improves routine diagnosis of extrapulmonary tuberculosis in a low-resource setting: A study from the tertiary care hospital in Zanzibar.

BACKGROUND: Extrapulmonary tuberculosis (EPTB) is a diagnostic challenge. An immunochemistry-based MPT64 antigen detection test (MPT64 test) has reported higher sensitivity in the diagnosis of EPTB compared with conventional methods. The objective of this study was to implement and evaluate the MPT64 test in routine diagnostics in a low-resource setting. METHODS: Patients with presumptive EPTB were prospectively enrolled at Mnazi Mmoja Hospital, Zanzibar, and followed to the end of treatment. Specimens collected were subjected to routine diagnostics, GeneXpert® MTB/RIF assay and the MPT64 test. The performance of the MPT64 test was assessed using a composite reference standard, defining the patients as tuberculosis (TB) cases or non-TB cases. RESULTS: Patients (n = 132) were classified as confirmed TB (n = 12), probable TB (n = 34), possible TB (n = 18), non-TB (n = 62) and uncategorized (n = 6) cases. Overall, in comparison to the composite reference standard for diagnosis, the sensitivity, specificity, positive predictive value, negative predictive value and accuracy of the MPT64 test was 69%, 95%, 94%, 75% and 82%, respectively. The MPT64 test performance was best in TB lymphadenitis cases (n = 67, sensitivity 79%, specificity 97%) and in paediatric TB (n = 41, sensitivity 100%, specificity 96%). CONCLUSIONS: We show that the MPT64 test can be implemented in routine diagnostics in a low-resource setting and improves the diagnosis of EPTB, especially in TB lymphadenitis and in children.