Multi-locus sequence typing of Escherichia coli isolated from clinical samples in Jordan.

INTRODUCTION: Escherichia coli (E. coli) is the major cause of extraintestinal infections in the urinary tracts and bloodstream in humans in the community and health care institutions. Several studies on the genetic characterization of E. coli among clinical and environmental isolates were performed and revealed a wide diversity of sequence types (STs). In Jordan, phenotypic and genetic features of E. coli were extensively studied but there is still a need to identify the STs that inhabit the community. METHODOLOGY: In this study, multi-locus sequence typing (MLST) was performed on archived clinical E. coli isolates collected from different hospitals in Jordan and the identified STs were extensively analyzed. RESULTS: Genotyping of 92 E. coli isolates revealed 34 STs and 9 clonal complexes. The frequencies of STs ranged between 1 to 23 observations. The most frequent STs among E. coli isolates were ST131 (n = 23), ST69 (n = 19), ST998 (n = 7), ST2083 (n = 5), and ST540 (n = 4). These five ST accounted for up to 60% of the 92 E. coli isolates. Based on the MLST database, the STs reported in this work were world widely recognized in humans, animals, and in the environment. CONCLUSIONS: This study has elaborated more knowledge about the genotypes of E. coli in Jordan, with recommendations for future studies to correlate its genotypes with virulence and resistance genes.

One-step, multiplex, dual-function oligonucleotide of loop-mediated isothermal amplification assay for the detection of pathogenic Burkholderia pseudomallei.

This study highlights the development of a multiplex real-time loop-mediated isothermal amplification assay. The developed assay employed a dual-function oligonucleotide (DFO) which simultaneously monitors the emitted amplification signals and accelerates the amplification process. The DFO was a modification of loop primer (LP); the 5'-end and 3'-end of the LP was tagged with fluorophore and quencher, respectively. The DFO was quenched in its unbound state and fluoresces only when it anneals to the specific target during the amplification process. With the same working mechanism as LP, DFO allowed the detection of target genes in less than 1 h in a real time monitoring system. We demonstrated this detection platform with Burkholderia pseudomallei, the causative agent of melioidosis. An internal amplification control (IAC) was incorporated in the assay to rule out false negative result and to demonstrate that the assay was successfully developed in a multiplex system. The assay was 100% specific when it was evaluated against 96 B. pseudomallei clinical isolates and 48 other bacteria species. The detection limit (sensitivity) of the developed assay was 1 fg/µl of B. pseudomallei genomic DNA and 18.2 CFU/ml at the bacterial cell level. In spiked blood samples, the assay's detection limit was 14 CFU/ml. The assay's diagnostic evaluation showed 100% diagnostic sensitivity, diagnostic specificity, positive predictive value, and negative predictive value. An integrated multiplex LAMP and real-time monitoring system was successfully developed, simplifying the workflow for the rapid and specific nucleic acid diagnostic test.

The status of cryptosporidiosis in Jordan: a review.

BACKGROUND: Cryptosporidium is a waterborne intestinal parasite that causes diarrhoea in low and middle-income countries worldwide. Reports from Mediterranean countries have documented the prevalence of cryptosporidiosis in children at various ages, also among cancer patients, and in cases of chronic kidney disease, haemodialysis, and organ transplant. Untill now, modified-acid staining preceded by stool concentration preparation remains the leading screening diagnostic test for the infection. In Jordan, few studies for cryptosporidiosis have been performed during the last 3 decades. AIMS: This paper reviewed the status of cryptosporidiosis in Jordan and tracked recent updates for this emerging protozoal infection among different population groups. METHODS: In this study, an online search was conducted on Google Scholar and PubMed databases using the keywords: Jordan, cryptosporidiosis and Cryptosporidium to inspect studies done on this parasite in Jordan. RESULTS: Only 9 articles were identified from 1994 to 2019. These were analysed in terms of population group, demography, clinical history and the diagnostic tools used. CONCLUSION: Cryptosporidiosis is still neglected in Jordan as indicated by the low number of studies over the last 3 decades and the prevalence is diverse depending on the diagnostic test used and socioeconomic status.

Cryptosporidiosis among Hemodialysis Patients in Jordan: First Preliminary Screening Surveillance.

Few studies have reported the incidence of cryptosporidiosis among hemodialysis patients worldwide. Currently many molecular and immunological assays have been developed for the sensitive diagnosis of cryptosporidiosis, but still, the microscopic detection of the parasitic infective stage (oocysts) in stool specimens using modified acid stain is regarded as a reliable sensitive technique which is widely used in many clinical labs. In the present study, a total of 133 stool samples were collected from hemodialysis patients and were screened for Cryptosporidium oocyst using formalin-ether concentration and modified acid-fast staining technique. Clinical and demographic data were also collected and analyzed. Cryptosporidium oocysts were recovered in 15/133 (11%) of the investigated hemodialysis patients. The age of patients ranged from 25 to 80 years (mean: 57.84 ± 12.22). Most of the Cryptosporidium-positive cases were recovered from males (73.7%) residing in rural villages in Irbid city (86.6%). The most repeatedly reported symptoms in the Cryptosporidium-positive patients were gastrointestinal symptoms, including diarrhea (15%), nausea (24%), abdominal pain (23%) and bloating (17%), in addition to general fatigue (32%) and weight loss (19%). No statistically significant associations for certain clinical symptoms or risk factors were found. The present study is the first preliminary study in Jordan that provided a brief screening for the incidence of cryptosporidiosis among hemodialysis patients.

Osteoarticular and soft-tissue melioidosis in Malaysia: clinical characteristics and molecular typing of the causative agent.

INTRODUCTION: Melioidosis involving bone, joints, and soft tissue is rare and reported usually following dissemination of disease from infection elsewhere in the body; to a lesser degree, it can also be reported as the primary manifestation of melioidosis. METHODOLOGY: The orthopedic registry at Hospital University Sains Malaysia from 2008 until 2014 was retrospectively reviewed and was followed by molecular typing of Burkholderia pseudomallei. RESULTS: Out of 20 cases identified, 19 patients were confirmed to have osteoarticular and/or soft-tissue melioidosis. The majority of the patients were males (84%), and 16 patients had underlying diabetes mellitus with no significant estimated risk with the disease outcomes. Bacterial genotype was not associated with the disease as a risk. Death was a significant outcome in patients with bacteremic infections (p = 0.044). CONCLUSION: Patients with lung or skin melioidosis require careful treatment follow-up to minimize the chance for secondary osteoarticular infection. Human risk factors remain the leading predisposing factors for melioidosis. Early laboratory and clinical diagnosis and acute-phase treatment can decrease morbidity and mortality.

The epidemiology and clinical spectrum of melioidosis in a teaching hospital in a North-Eastern state of Malaysia: a fifteen-year review.

BACKGROUND: Over the last two decades, many epidemiological studies were performed to describe risks and clinical presentations of melioidosis in endemic countries. METHODS: We performed a retrospective analysis of 158 confirmed cases of melioidosis collected from medical records from 2001 to 2015 in Hospital Universiti Sains Malaysia, Kubang Kerian, Kelantan, Malaysia, in order to update the current status of melioidosis clinical epidemiology in this putatively high risk region of the country. RESULTS: Principal presentations in patients were lung infection in 65 (41.1 %), skin infection in 44 (27.8 %), septic arthritis/osteomyelitis in 20 (12.7 %) and liver infection in 19 (12.0 %). Bacteremic melioidosis was seen in most of patients (n = 121, 76.6 %). Focal melioidosis was seen in 124 (78.5 %) of patients and multi-focal melioidosis was reported in 45 (28.5 %) cases. Melioidosis with no evident focus was in 34 (21.5 %) patients. Fifty-four (34.2 %) patients developed septic shock. Internal organ abscesses and secondary foci in lungs and/or soft tissue were common. A total of 67 (41 %) cases presented during the monsoonal wet season. Death due to melioidosis was reported in 52 (32.9 %) patients, while relapses were occurred in 11 (7.0 %). Twelve fatal melioidosis cases seen in this study were directly attributed to the absence of prompt acute-phase treatment. Predisposing risk factors were reported in most of patients (n = 133, 84.2 %) and included diabetes (74.7 %), immune disturbances (9.5 %), cancer (4.4 %) and chronic kidney disease (11.4 %). On multivariate analysis, the only independent predictors of mortality were the presence of at least one co-morbid factor (OR 3.0; 95 % CI 1.1-8.4), the happening of septic shock (OR 16.5; 95 % CI 6.1-44.9) and age > 40 years (OR 6.47; 95 % CI 1.7-23.8). CONCLUSIONS: Melioidosis should be recognized as an opportunistic nonfatal infection for healthy person. Prompt early diagnosis and appropriate antibiotics administration and critical care help in improved management and minimizing risks for death.

Brief communication genotyping of Burkholderia pseudomallei revealed high genetic variability among isolates from a single population group.

Burkholderia pseudomallei is a soil dwelling Gram-negative bacteria predominates in Southeast Asia zone and the tropical part of Australia. Genetic diversity has been explored among various populations and environments worldwide. To date, little data is available on MLST profiling of clinical B. pseudomallei isolates in peninsular Malaysia. In this brief report, thirteen culture positive B. pseudomallei cases collected from a single population of Terengganu state in the Western Peninsular Malaysia and were confirmed by In-house TTS1-PCR. Isolates were subjected for multi-locus sequence typing (MLST) to explore their genotypic diversity and to investigate for possible clonal clustering of a certain sequence type. Patient's clinical information was examined to investigate for clinical correlation among the different genotypes. In spite of small sample set, MLST results indicated predictive results; considerable genotypic diversity, predominance and novelty among B. pseudomallei collected over a single geographically-located population in Malaysia. Massive genotypic heterogeneity was observed; 8 different sequence types with predominance of sequence type 54 and discovery of two novel sequence types. However, no clear pathogenomic or organ tropism clonal relationships were predicted.

Detection of Strongyloides stercoralis infection among cancer patients in a major hospital in Kelantan, Malaysia.

INTRODUCTION: Strongyloidiasis is one of the most commonly neglected but clinically important parasitic infections worldwide, especially among immunocompromised patients. Evidence of infection among immunocompromised patients in Malaysia is, however, lacking. In this study, microscopy, real-time polymerase chain reaction (PCR) and enzyme-linked immunosorbent assays (ELISAs) were used to detect Strongyloides stercoralis (S. stercoralis) infection among cancer patients in a Malaysian hospital. METHODS: A total of 192 stool and serum samples were collected from cancer patients who were receiving chemotherapy with or without steroid treatment at a hospital in northeastern Malaysia. Stool samples were examined for S. stercoralis using parasitological methods and real-time PCR. Serology by ELISA was performed to detect parasite-specific immunoglobulin G (IgG), IgG4 and immunoglobulin E (IgE) antibodies. For comparison, IgG4- and IgG-ELISAs were also performed on the sera of 150 healthy individuals from the same area. RESULTS: Of the 192 samples examined, 1 (0.5%) sample was positive for S. stercoralis by microscopy, 3 (1.6%) by real-time PCR, 8 (4.2%) by IgG-ELISA, 6 (3.1%) by IgG4-ELISA, and none was positive by IgE-ELISA. In comparison, healthy blood donors had significantly lower prevalence of parasite-specific IgG (2.67%, p < 0.05) and IgG4 (2.67%, p < 0.05) responses. CONCLUSION: This study showed that laboratory testing may be considered as a diagnostic investigation for S. stercoralis among immunocompromised cancer patients.