Molecular and serological detection of Leptospira and Ricktettsia in patients with acute febrile illness

Leptospirosis is a zoonotic disease which is caused by the spirochete, Leptospira interrogans (L interrogans). Rodents generally serve as the main carrier for Leptospirosis and transmit the disease to humans through cuts and abrasions in the skin or through mucous membranes of the eyes, nose or throat. High seroprevalence was recorded in Sabah, among people living within the national park is likely due to high exposure and contact with wild animals. Diagnosis of Leptospirosis is challenging due to a wide diversity of clinical symptoms which mimic regular symptoms of fever. Lipl32 is a major outer membrane protein, which is present only in pathogenic strains of Leptospira. Studies show that ELISA is able to detect Leptospira-specific antibodies earlier than the gold standard method, the Microscopic Agglutination Test (MAT). In this study, we have used codon optimized synthetic gene encoding Lipl32 and transfected into E coli expression vector BL21 (DE3). The recombinant Lipl32 protein was expressed after induction with IPTG which resulted in approximately a 40kDa protein. The purified recombinant Lipl32 protein was used as an antigen for detecting Leptospira-specific antibody by ELISA. Preliminary_ results showed that the recombinant Lipl32 antigen was able to detect Leptospira­specific IgM and IgG in human serum samples. Hence, this method could be used for diagnostic purposes and in epidemiological investigations of Leptospirosis

Characterization of the pscC (Type III secretion) gene of Pseudomonas aeruginosa (PA01) and assessment of immunogenicity of pscC protein in rats

Proteins associated with the bacterial membrane can be recruited for application as antigens for the development of vaccines. This preliminary study was directed towards evaluating the antigenic properties of the Pseudomonas aeruginosa (PA01) pscC protein which is a component of the Type III secretion system. Gene specific primers were designed to isolate the pscC gene which was isolated, ligated onto the multiple cloning site of vector pGS21(a), cloned and expressed in Escherichia coli (BL21). The molecular weight of the expressed pscC protein was determined by SDS-PAGE (10% sodium dodecyl sulphate-polyacrylamide gel electrophoresis) and was found to be around 57 KDa and purified by the size exclusion chromatography. Finally, the purified pscC protein was injected subcutaneously into adult Sprague Dawley rats with a range of concentrations (50, 100 and 150 microgram per rat) respectively. Recombinant pscC antigen induced a specific humoral immune response against the antigen, which was validated by Enzyme-linked immunosorbent assay (ELISA). The results concluded that anti-pscC antibody was elicited in the animal model

Molecular characteristics of infection and colonization isolates of community-acquired methicillin-resistant Staphylococcus aureus (CA-MRSA)

Staphylococcus aureus is a gram-positive coccus that colonizes the skin and mucous membranes, particularly the anterior nares. Recently, community-acquired MRSA (CA-MRSA) has emerged as a cause of skin and soft-tissue infections in healthy individuals. These strains are sensitive to antimicrobials, carry genes for Panton-Valentine leukocidin (PVL) toxin, and feature the staphylococcal cassette chromosome mec (SCCmec) type IV or V. The suspected mode of transmission involves close contact with carriers, leading to skin or nasal colonization that results in subsequent active infection. This study was undertaken to determine the molecular characteristics of CA-MRSA isolates in children presenting with wound infections at Likas Hospital, Sabah, Malaysia, and the possible mode of transmission. The results showed that the majority of CA-MRSA infection isolates were from scalp abscesses (49%) in 1–5-year-old children (70%) in the Filipino (54%) community. The presence of the mec gene was detected in all isolates and the PVL virulence factor was found in 92% of the isolates. SCCmec typing revealed that 57% of the isolates were untypable, 35% harbored the SCCmecIVa element, and one each had SCCmecIVc, SCCmecV, or SCCmecII. Sixteen S. aureus strains were isolated from nasal swabs in 19 family members of index patients. Fourteen of these cultures were positive for catalase, coagulase, and DNAase. All of the colonization isolates carried the mecA gene and only a third were positive for the PVL toxin. SCCmec typing showed that 79% of the isolates were untypable and two had SCCmecIVa element and one had SCCmecV element. When five pairs of infection and colonizing isolates were compared by spa typing, only two pairs showed identical spa type with possible transmission between the patient and family contact. Further studies are necessary to establish CA-MRSA transmission by performing multiple-site cultures multiple times instead of one-time naresonly sample collection

Recombinant LipL32 protein developed using a synthetic gene detects leptospira-specific antibodies in human serum samples

Background: Synthetic biology is emerging as a viable alternative for the production of recombinant antigens for diagnostic applications. It offers a safe alternative for the synthesis of antigenic principles derived from organisms that pose a high biological risk. Methods: Here, we describe an enzyme-linked immunosorbent assay (ELISA) using the synthetic recombinant LipL32 (rLipL32) protein expressed in Escherichia coli for the detection of Leptospira-specific antibodies in human serum samples. The rLipL32-based ELISA was compared with a microscopic agglutination test (MAT), which is currently used as the gold standard for the diagnosis of leptospirosis. Results: Our results showed that all the MAT-positive serum samples were positive for Leptospira-specific IgG in an ELISA, while 65% (n = 13) of these samples were also positive for Leptospira-specific IgM. In the MAT-negative serum samples, 80% and 55% of the samples were detected as negative by an ELISA for Leptospira-specific IgM and IgG, respectively. Conclusion: An ELISA using the synthetic rLipL32 antigen was able to distinguish Leptospira-specific IgM (sensitivity 65% and specificity 80%) and IgG (sensitivity 100% and specificity 55%) in human serum samples and has the potential to serve as a rapid diagnostic test for leptospirosis

Brain Abscess in the Current Decade (2010–2019) in India—A Review

Brain abscess outcomes have improved in recent years due to advancements in cranial imaging, microbiological techniques, minimally invasive neurosurgical procedures, and effective antibiotic treatments. However, the incidence of brain abscess remains unchanged in developing countries. We searched PubMed and Google Scholar for references using the key words “brain abscess” and “India” and reviewed both retrospective and prospective studies published in peer-reviewed journals in the current decade to understand the present status. The review shows that the patients’ ages, the predominance of male patients, the symptoms and locations of brain abscesses, and the types of bacteria associated with them have remained unchanged over the past decade. The most common predisposing condition in recent years has been chronic suppurative otitis media with a mortality rate of 7 to 10%. Middle ear infection is often neglected and not treated aggressively in Asian countries. It requires multidisciplinary treatment strategies to address the primary source of infection and better health awareness to prevent the development of brain abscess

Whole genome sequencing of Mycobacterium tuberculosis SB24 isolated from Sabah, Malaysia

Mycobacterium tuberculosis (M. tuberculosis) is the causative agent of tuberculosis (TB) that causes millions of death every year. We have sequenced the genome of M. tuberculosis isolated from cerebrospinal fluid (CSF) of a patient diagnosed with tuberculous meningitis (TBM). The isolated strain was referred as M. tuberculosis SB24. Genomic DNA of the M. tuberculosis SB24 was extracted and subjected to whole genome sequencing using PacBio platform. The draft genome size of M. tuberculosis SB24 was determined to be 4,452,489 bp with a G + C content of 65.6%. The whole genome shotgun project has been deposited in NCBI SRA under the accession number SRP076503

Expression distribution of cancer stem cells, epithelial to mesenchymal transition, and telomerase activity in breast cancer and their association with clinicopathologic characteristics

A total of 167 surgically resected primary invasive breast carcinomas and 63 metastatic lymph node lesions were analyzed for immunohistochemical (IHC) localization of the CD44+CD24−low breast cancer stem cell (CSC) markers, epithelial to mesenchymal transition (EMT) markers, and telomerase activity by double-staining IHC technique, in formalin-fixed, paraffin-embedded tissue, the results were validated by double-staining immunofluorescent and flow cytometry techniques. The results showed that CSCs with CD44+CD24−low phenotype were significantly increased in node-positive tumors, high-grade tumors, and ductal carcinoma in situ (DCIS). There was a high incidence of telomerase expression in metastatic lymph node lesion. There were considerably high number of tumor cells with EMT expression in metastatic lymph node lesion, and triple-negative tumor. The occurrence of EMT phenomena was usually accompanied by the co-existence of CSCs of CD44+CD24−low phenotype. There was no association between the existence of CSCs and detection of telomerase activity in tumor cells. Increased numbers of both CSCs of CD44+CD24−low phenotype and cells under-went EMT in DCIS lesion might be an initial step in the stromal invasion and propagation of breast cancer, and occurrence of EMT in the breast tumor associated with high prevalence of CSCs, promoting tumor invasiveness and metastasis

Metagenomic data of bacterial community from different land uses at the river basin, Kelantan

The data provided in the article includes the sequence of bacterial 16S rRNA gene from a high conservation value forest, logged forest, rubber plantation and oil palm plantation collected at Kelantan river basin. The logged forest area was previously notified as a flooding region. The total gDNA of bacterial community was amplified via polymerase chain reaction at V3-V4 regions using a pair of specific universal primer. Amplicons were sequenced on Illumina HiSeq paired-end platform to generate 250 bp paired-end raw reads. Several bioinformatics tools such as FLASH, QIIME and UPARSE were used to process the reads generated for OTU analysis. Meanwhile, R&D software was used to construct the taxonomy tree for all samples. Raw data files are available at the Sequence Read Archive (SRA), NCBI and data information can be found at the BioProject and BioSample, NCBI. The data shows the comparison of bacterial community between the natural forest and different land uses