Mochammad Hatta

Phylogeographical analysis of the dominant multidrug-resistant H58 clade of Salmonella Typhi identifies inter- and intracontinental transmission events

The emergence of multidrug-resistant (MDR) typhoid is a major global health threat affecting many countries where the disease is endemic. Here whole-genome sequence analysis of 1,832 Salmonella enterica serovar Typhi (S. Typhi) identifies a single dominant MDR lineage, H58, that has emerged and spread throughout Asia and Africa over the last 30 years. Our analysis identifies numerous transmissions of H58, including multiple transfers from Asia to Africa and an ongoing, unrecognized MDR epidemic within Africa itself. Notably, our analysis indicates that H58 lineages are displacing antibiotic-sensitive isolates, transforming the global population structure of this pathogen. H58 isolates can harbor a complex MDR element residing either on transmissible IncHI1 plasmids or within multiple chromosomal integration sites. We also identify new mutations that define the H58 lineage. This phylogeographical analysis provides a framework to facilitate global management of MDR typhoid and is applicable to similar MDR lineages emerging in other bacterial species.

Serum levels of interferon-γ, tumour necrosis factor-α, soluble interleukin-6R and soluble cell activation markers for monitoring response to treatment of leprosy reactions

Identifying pathogen and host‐related laboratory parameters are essential for the early diagnosis of leprosy reactions. The present study aimed to clarify the validity of measuring the profiles of serum cytokines [interleukin (IL)‐4, IL‐6, IL‐10, interferon (IFN)‐γ and tumour necrosis factor (TNF)‐α], the soluble IL‐6 receptor (sIL‐6R), soluble T cell (sCD27) and macrophage (neopterin) activation markers and Mycobacterium leprae‐specific anti‐PGL‐I IgM antibodies in relation to the leprosy spectrum and reactions. Serum samples from 131 Indonesian leprosy patients (82 non‐reactional leprosy patients and 49 reactional) and 112 healthy controls (HC) from the same endemic region were investigated. Forty‐four (89·8%) of the reactional patients had erythema nodosum leprosum (ENL) while only five (10·2%) had reversal reaction (RR). Follow‐up serum samples after corticosteroid treatment were also obtained from 17 of the patients with ENL and one with RR. A wide variability in cytokine levels was observed in the patient groups. However, IFN‐γ and sIL‐6R were elevated significantly in ENL compared to non‐ENL patients. Levels of IFN‐γ, TNF‐α and sIL‐6R declined significantly upon corticosteroid treatment of ENL. Thus, although the present study suggests limited applicability of serial measurement of IFN‐γ, TNF‐α and sIL‐6R in monitoring treatment efficacy of ENL, reactions it recommends a search for a wider panel of more disease‐specific markers in future studies.

Simple, rapid, and affordable point-of-care test for the serodiagnosis of typhoid fever

We developed a point-of-care test for the serodiagnosis of typhoid fever in the format of an immunochromatographic lateral flow assay. The flow assay for typhoid fever is based on the detection of Salmonella enterica serotype Typhi lipopolysaccharide-specific immunoglobulin M (IgM) antibodies. The assay was evaluated on serum samples collected in a hospital in South Sulawesi, Indonesia, where typhoid fever is endemic, and the results were compared with culture and Widal test. The sensitivity of this typhoid fever IgM flow assay for samples collected at 1st diagnosis from patients with culture-confirmed typhoid fever was determined to be 59.3%. The sensitivity ranged from 41.2% to 89.5%, depending on the duration of illness. A specificity of 97.8% was calculated based on results obtained for patients with clinical suspicion of typhoid fever that was later excluded. The assay is ideal for use as a point-of-care test in health care centers that lack the expertise and facilities to perform culture or the less specific Widal test. Because of its simplicity, the assay may also be used as a field test in remote areas.

Epidemiology of leprosy on five isolated islands in the Flores Sea, Indonesia.

We conducted a population‐based survey on five small islands in South Sulawesi Province (Indonesia) to collect baseline data previous to a chemoprophylactic intervention study aiming at interrupting the transmission of Mycobacterium leprae. Here we describe the present leprosy epidemiology on these geographically isolated islands. Of the 4774 inhabitants living in the study area 4140 were screened for leprosy (coverage: 87%). We identified 96 leprosy patients (85 new and 11 old patients), representing a new case detection rate (CDR) of 205/10 000 and a prevalence rate of 195/10 000. CDRs were similar for males and females. Male patients were more often classified as multibacillary (MB) than women. Of the new patients, 33 (39%) were classified as MB, 16 (19%) as paucibacillary (PB) 2–5 lesions and 36 (42%) as PB single lesion. In this area of high leprosy endemicity leprosy patients were extensively clustered, i.e. not equally distributed among the islands and within the islands among the houses.

Genetic, household and spatial clustering of leprosy on an island in Indonesia: a population-based study

BackgroundIt is generally accepted that genetic factors play a role in susceptibility to both leprosy per se and leprosy type, but only few studies have tempted to quantify this. Estimating the contribution of genetic factors to clustering of leprosy within families is difficult since these persons often share the same environment. The first aim of this study was to test which correlation structure (genetic, household or spatial) gives the best explanation for the distribution of leprosy patients and seropositive persons and second to quantify the role of genetic factors in the occurrence of leprosy and seropositivity.MethodsThe three correlation structures were proposed for population data (n = 560), collected on a geographically isolated island highly endemic for leprosy, to explain the distribution of leprosy per se, leprosy type and persons harbouring Mycobacterium leprae-specific antibodies. Heritability estimates and risk ratios for siblings were calculated to quantify the genetic effect. Leprosy was clinically diagnosed and specific anti-M. leprae antibodies were measured using ELISA.ResultsFor leprosy per se in the total population the genetic correlation structure fitted best. In the population with relative stable household status (persons under 21 years and above 39 years) all structures were significant. For multibacillary leprosy (MB) genetic factors seemed more important than for paucibacillary leprosy. Seropositivity could be explained best by the spatial model, but the genetic model was also significant. Heritability was 57% for leprosy per se and 31% for seropositivity.ConclusionGenetic factors seem to play an important role in the clustering of patients with a more advanced form of leprosy, and they could explain more than half of the total phenotypic variance.

Induction of roots on jujube softwood cuttings using Agrobacterium rhizogenes

SummaryCuttings from trees of jujube (Ziziphus jujuba Mill.), particularly mature ones, root with great difficulty. A greenhouse experiment assessed the effects of two strains of Agrobacterium rhizogenes (A4 and TR105) on softwood cuttings from two sources: ten year old trees of cv. Li and a tree approximately 40 years old of the cv. Contorta. The cuttings were collected and trimmed to 7.5 cm. Ten weeks after inoculation, both strain and source of cutting influenced visible symptoms of infection. Jujube cuttings were more responsive to TR105 than to A4, and the ‘Li’ cuttings were more responsive than ‘Contorta’ cuttings. Strain TR105 was very effective in increasing rooting percentages and root number. ‘Li’ cuttings had better rooting percentages, greater root number, and greater root length than did the ‘Contorta’ cuttings. Inoculation with A. rhizogenes may also possess great potential for promoting adventitious root formation in other difficult woody ornamental or fruit tree species.

An extended genotyping framework for Salmonella enterica serovar Typhi, the cause of human typhoid

The population of Salmonella enterica serovar Typhi (S. Typhi), the causative agent of typhoid fever, exhibits limited DNA sequence variation, which complicates efforts to rationally discriminate individual isolates. Here we utilize data from whole-genome sequences (WGS) of nearly 2,000 isolates sourced from over 60 countries to generate a robust genotyping scheme that is phylogenetically informative and compatible with a range of assays. These data show that, with the exception of the rapidly disseminating H58 subclade (now designated genotype 4.3.1), the global S. Typhi population is highly structured and includes dozens of subclades that display geographical restriction. The genotyping approach presented here can be used to interrogate local S. Typhi populations and help identify recent introductions of S. Typhi into new or previously endemic locations, providing information on their likely geographical source. This approach can be used to classify clinical isolates and provides a universal framework for further experimental investigations.

Increased chitotriosidase activity in serum of leprosy patients: Association with bacillary leprosy

Human phagocyte-specific chitotriosidase is associated with several diseases involving macrophage activation. Since macrophage activation plays an important role in the control of Mycobacterium leprae infection, we studied the association of chitotriosidase with leprosy both in serum and in situ in lesional skin biopsies from patients. Serum samples from 78 Indonesian leprosy patients (39 non-reactional and 39 reactional leprosy patients) and 36 healthy controls (HC) from the same endemic region were investigated. The patients were classified as multibacillary (MB, n=69) or paucibacillary (PB, n=9) based on the bacterial index in slit-skin smears. Thirty-six of the reactional patients had erythema nodosum leprosum (ENL), while only 3 had reversal reaction (RR). Follow-up serum samples after corticosteroid treatment were also obtained from 17 patients with ENL and one with RR. Multibacillary (MB) patients showed increased chitotriosidase activity in serum as compared to paucibacillary (PB) patients and healthy controls. Although no significant difference was observed between reactional and the corresponding non-reactional groups, ENL showed significantly higher chitotriosidase activity as compared to HC. Furthermore, corticosteroid treatment resulted in significant decline of enzyme activity in ENL sera. Chitotriosidase activity correlated with levels of neopterin, another macrophage activation marker, but not with IL-6, IFN-gamma, TNF-alpha and IL-10. Immunohistochemical staining of 6 MB (LL=5, BL=1) lesional skin sections from stored material showed positive staining for chitotriosidase within lipid-laden macrophages suggesting that macrophages are the source of the enzyme detected in serum. Thus, serum chitotriosidase activity is potentially useful in distinguishing MB from PB leprosy and in monitoring response to therapy in ENL.

Detection and identification of mycobacteria in sputum from suspected tuberculosis patients

BackgroundDetection of Tuberculosis agent like nontuberculous mycobacteria (NTM) species by culture and microscopic methods remains difficult and time consuming. A fast and reliable diagnosis of tuberculosis would greatly improve the control of the disease. The purpose of this study is to compare the conventional multiplex PCR and multiplex PCR reverse cross blot hybridization assay to culture method in terms of mycobacteria species detection.FindingsAmong the 117 positively cultured samples, nontuberculous mycobacteria (NTM) species were found in 9 samples of multiplex PCR reverse cross blot hybridization assay; compared to only 3 NTM species found in our conventional multiplex PCR, and 13 NTM species were successfully identified among 162 negatively cultured samples compared to only 5 NTM species identification in conventional multiplex PCR results.ConclusionsThe sensitivity of the multiplex PCR reverse cross blot hybridization assay comparing to culture method was 86.03%, the specificity is 35.46%, the positive predictive value was 41.94% and the negative predictive value was 82.41%. For conventional multiplex PCR these values are 81.62%, 38.65%, 41.89%, 79.51%, respectively. Furthermore, in terms of mycobacteria species detection, the conventional multiplex PCR was relatively equal compared to the multiplex PCR reverse cross blot hybridization assay, and to be particularly having no significant discrepant results on the identification of Mycobacteria tuberculosis in both methods.

Brucellosis seroprevalence in Bali cattle with reproductive failure in South Sulawesi and Brucella abortus biovar 1 genotypes in the Eastern Indonesian archipelago.

BackgroundBrucellosis is a major cause of infertility and reproductive failure in livestock. While cattle in the Eastern Indonesian archipelago suffers from reproductive problems information on bovine brucellosis in the region is fragmentary. The control of brucellosis requires a major and prolonged effort and confirmation of the infection by isolation with detailed knowledge of the spread of the infection is essential when planning a control program.ResultsSerological investigation of Brucella infection in beef cattle tended under extensive farming conditions revealed a high seroprevalence (19.3%; 95% CI, 17–22) in the compliment fixation tests. The results of a rapid and simple field test correlated well with the Rose Bengal test (kappa, 0.917) and indicated an acceptable sensitivity (87.5%) and specificity (98.1%) compared with the complement fixation test. Reproductive failure was reported for 39.0% of the cows with a loss of calves due to abortion or early death amounting to 19.3%. Past reproductive failure did not, however, correlate with seropositivity in the complement fixation test (RP = 1.21; P = 0.847). B. abortus biovar 1 was freshly isolated from the hygromas of two cows and together with thirty banked isolates collected since 1990 from different parts of Sulawesi and Timor eight related genotypes could be distinguished with one genotype being identical to that of an isolate (BfR91) from Switzerland. The Indonesian genotypes formed together with BfR91 and one African and one North American isolate a distinct branch on the B. abortus biovar 1 dendogram.ConclusionsBovine brucellosis appears to be widespread in the Eastern Indonesian archipelago and calls for urgent intervention. The fresh isolation of the pathogen together with the observed high seroprevalence demonstrates the presence and frequent exposure of cattle in the area to the pathogen. The application of a rapid and simple field test for brucellosis could be very useful for the quick screening of cattle at the pen side.