Potential of Colicin as an Antibacterial Agent in Escherichia coli

The development of antibiotics calls for the critical consideration of instances of resistance. Infectious disorders brought on by resistant bacterial infections could affect the entire world. It is believed that the protein that the bacteria generate may one day replace antibiotics as an alternative antibacterial agent. Both Gram-positive and Gram-negative bacteria have the ability to manufacture bacteriocin. The bacteriocin type produced by Escherichia coli, notably colicin, has been demonstrated to inhibit the same bacteria through various essential methods. Colicin, a substance made by an E. coli cell, is also capable of protecting itself from attack; however, this defense mechanism has not yet been identified. The traits of colicin and the method by which it functions as a different antimicrobial agent to inhibit other bacteria will be covered in this article. We analyze the potential of colicin as an antibacterial agent in E. coli using PRISMA methods from diverse academic sources. Here, we found that the structure of the colicin, namely its central receptor domain, aids in the recognition of target cells. Promising results were found in recent studies on the antibacterial effects of the E. coli and colicin combinatio

Evaluation of Tuberculosis Vaccine Candidate, pcDNA3.1-rpfD using Mycobacterial Growth Inhibition Assay (MGIA)

Resuscitation-promoting factor D (RpfD) is a protein involved in the resuscitation of dormant bacteria. A new tuberculosis vaccine carrying the rpfD gene has been successfully constructed, pcDNA3.1-rpfD. It was demonstrated that this vaccine exhibits cellular and humoral immune responses. Therefore, within this study, the efficacy of this new vaccine candidate was evaluated using mycobacterial growth inhibition assay (MGIA). MGIA is a functional assay that measures the complex host immune response, peripheral blood mononuclear cell (PBMC) and splenocyte from BALB/c mice against mycobacteria. With BACTECTM MGITTM 960 automated system, the effect of vaccination on bacterial growth was reported as a time to positivity (TTP) in hours. The mean of TTP from the vaccinated group (both pcDNA3.1-rpfD and BCG) was higher than the negative control group. These results suggest that pcDNA3.1-rpfD may be effective in controlling tuberculosis growth and may provide a clue for the development of the tuberculosis vaccine. In addition, despite previous evidence that IFNγ was essential for tuberculosis immunity, IFNγ (interferon gamma) production was found not to be correlated with mycobacterial inhibition. Therefore, these findings offer an alternative method to evaluate vaccine candidates than the assessment using IFNγ only

The Existence of Mycobacterium tuberculosis in Microenvironment of Bone

Mycobacterium tuberculosis is an obligate aerobe bacteria requiring oxygen in its metabolism. In normal condition, bones have pH of 6.9–7.4 and temperature of 37°C. With the composition mentioned, bones fall in the group of tissue with less rich oxygen (<35%) which theoretically means, M. tuberculosis is hard to grow in the bone environment. Bone microliving environment is formed by the cells constructing the bone itself and the active cells which periodically interact with the bone cells. Activation of these cells gives impact to the temperature, pH, gas concentration, and liquid concentration, and at the same time triggers calcium, phosphor, and other minerals to be deposited in the bone. In the process of new bone formation, the osteoblast cells produce matrix and release them to the microenvironment that needs a high concentration of calcium and phosphor. The survival of M. tuberculosis in the microenvironment of bone is reflected in interaction of the bacteria and the non-immune cells, the bacteria and the organic environment, and the bacteria and the inorganic environment. In addition, the immune system also threatens the survival of M. tuberculosis. The results of these interactions will affect the lives of bacteria and has an impact on the bone microenvironment

CLONING AND EXPRESSION OF MCE1A GENE FROM MYCOBACTERIUM TUBERCULOSIS BEIJING AND H37RV STRAIN FOR VACCINE CANDIDATE DEVELOPMENT

Background: Tuberculosis remains the leading cause of death in the world, especially wherever poverty, malnutrition and poor housing prevail. Mycobacterium tuberculosis Beijing strain is the most common strain that causes tuberculosis in Indonesia. The wide spread of tuberculosis has been further aggravated by HIV-AIDS and drug resistance. Unfortunately, Bacille Calmette-Guerin (BCG) as the current vaccine has different protection function and efficacy. According to function analysis, mce1A gene was predicted to have a role in host invasion and survival of Mycobacterium tuberculosis in human macrophages. Materials and Methods: We performed cloning and protein expression of Mce1A gene of Mycobacterium tuberculosis Beijing strain as local isolate and standard strain H37Rv as a comparison on the expression system Escherichia coli BL21(DE3). Mce1A gene from the strains were amplified by PCR and inserted into the vector pET28a. Each resulting recombinant plasmid was subsequently transformed into E. coli BL21(DE3) and Mce1A protein was expressed with IPTG induction. Results: E. coli BL21(DE3) was succesfully transformed with a recombinant plasmid that contains the Mce1A gene insert with correct orientation and reading frame. There was no mutation found in the amino acids sequence for B and T cell epitope. Mce1A expression in E. coli BL21(DE3) showed a protein band that was higher than expected. The protein was confirmed with Western blotting using anti-His detector. Conclusion: We assumed that Mce1A recombinant protein that has been expressed in E. coli BL21(DE3) is in their dimeric form or alternatively formed aggregates of different sizes

Acinetobacter baumannii

Acinetobacter baumannii (A. baumannii) is Gram-negative coccobacilli that has emerged as a nosocomial pathogen. Several reports in Indonesia showed the continuous presence of A. baumannii. This study aimed to determine the incidence of A. baumannii bacteremia in neonates in the Neonatal Unit Dr. Cipto Mangunkusumo Hospital (RSCM), Jakarta, Indonesia, and assess its role in blood stream infection using antibiogram and genotyping by pulsed-field gel electrophoresis (PFGE). Subjects were neonates with clinical sepsis. Blood specimens from the neonates and samples of suspected environment within the Neonatal Unit were cultivated. Antimicrobial resistance profiles were classified for analysis purpose. A. baumannii isolates were genotyped by PFGE to determine their similarity. A total of 24 A. baumannii were isolated from 80 neonates and the environment during this period of study. Seven isolates from the neonates showed multiple antimicrobial resistance (MDR), and 82% (n=17) of the environment isolates were also MDR. Antibiotype “d” seemed to be predominant (62.5%). PFGE analysis showed a very close genetic relationship between the patients and environment isolates (Dice coefficient 0.8–1.0). We concluded that a mode of transmission of environmental microbes to patients was present in the Neonatal Unit of RSCM and thus needed to be overcome

A comparison study of GeneXpert and In-House N1N2 CDC Real-Time RT-PCR for detection of SARS-CoV-2 infection

COVID-19 is a disease caused by SARS-CoV-2, a new virus from genus β-coronaviruses. This disease has been declared a pandemic by WHO on 11 March 2020 until now. The nucleic acid tests are the most frequently used assays because of their high sensitivity and specificity. One of the tests is the GeneXpert, a real-time reverse transcription polymerase chain reaction (rRT-PCR)-based assay platform. The use of the GeneXpert shows great public health interest because of the rapid (50 min), the minimum number of trained staff, and less infrastructure and equipment. However, there are limited data on the application of the GeneXpert for the detection of SARS-CoV-2. Therefore, we conducted a comparative study between the GeneXpert and in-house N1N2 CDC rRT-PCR assay. Of 86 samples, 17 were rRT-PCR positive while 13 were GeneXpert positive. Of rRT-PCR positive 17 samples, 7 were GeneXpert negative [58.82% (10/17] sensitivity]. We also found that 3 GeneXpert positive samples showed rRT-PCR negative (95.65% [66/69] specificity). It is concluded that negative results by the GeneXpert can not rule out the possibility of SARS-CoV-2 infection, particularly in close-contact individuals and the interpretation of the positive result should be analyzed carefully, particularly amplification with Ct>40

Integration of water, sanitation, and hygiene program with biosecurity: A One Health approach to reduce the prevalence and exposure of antibiotic-resistant bacteria in the livestock community

The global spread of antibiotic resistance poses a significant threat to public health and is one of the main causes of this problem. Livestock farming plays a significant role in the horizontal and vertical transmission of treatment-resistant genes and bacteria. These processes involve contact with agricultural products and the environment, raising concerns for public health, and farming communities. The farming community is composed of a staggering 608 million farms worldwide, and their livelihood depends heavily on livestock farming. To address this issue, a multidisciplinary One Health approach focusing on integrated monitoring and intervention for humans, animals, and the environment is essential. Water, sanitation, and hygiene (WaSH) programs have the potential to significantly reduce the risk of exposure to antibiotic-resistant bacteria, particularly extended spectrum beta-lactamase (ESBL) Escherichia coli, by obstructing the transmission route between humans and animals. Additional risk reduction measures for ESBL E. coli infection in animals include vaccination and biosecurity program implementation. Water, sanitation, and hygiene and biosecurity measures must be combined to maximize the effectiveness of the One Health program. Therefore, this study aimed to describe recent advances in biosecurity and WaSH interventions in the livestock environment, analyze the effects of these interventions on human and animal health, and investigate potential future scenarios within the quantitative microbial risk assessment framework. This study used an integrative literature review through searches of four databases, a review of World Health Organization documents through websites, and an examination of relevant texts from previously obtained reference lists. Although hygiene and sanitation are often combined, there is still a lack of quantitative evaluation of the efficacy of integrating WaSH with biosecurity in livestock. In addition, the integration of the WaSH program with biosecurity has potential as a One Health intervention in the coming years

サイボウヘキ ゴウセイ オ ソガイスル コウセイ ブッシツ エンヂュラシジン ニ タイスル コソウキン ノ テンシャ オウトウ ノ タイリング チップ ニ ヨル ヒョウカ

博士(Doctor)バイオサイエンス(Bioscience)奈良先端科学技術大学院大学博第882号乙第882号博士(バイオサイエンス)奈良先端科学技術大学院大

Optimization of pGEX System to Express and Isolate Mycobacterium tuberculosis Inclusion Body Protein in Combining with Modified Refolding Method

Antigen sub units for vaccine studies are typically isolated from recombinant proteins in an expression system. However, not all protein expression systems are used to express the specific protein. In this study, we optimized the pGEX system combined with the modified protein refolding to express and isolate M. tuberculosis proteins, especially proteins that are expressed as an inclusion body. Resuscitation promoting factor B (RpfB) protein is one of the Resuscitation promoting factor (Rpf) family of proteins that has been studied for its ability to induce cellular immunity in animal tests. Silico analyses demonstrate how RpfB is included in cell wall and cell processes. The Rpf family proteins are promising antigens that can be used as a TB vaccine candidate. The polymerase chain reaction was briefly performed using specific primers to amplify the full length of the rpfB. PCR amplification products were then purified, cut by restriction endonucleases, and cloned in to pGEX 6-P1. Protein expression was done in the Escherichia coli BL21 strain, and expressed protein was isolated using the modified protein refolding and solubilization method. The complex protein expression that appeared as inclusion bodies were successfully isolated and can be detected as complex GST-RpfB through the western blotting process. Our study results indicate that this system and our modified method are suitable for M. tuberculosis inclusion body protein expression and isolation