Exposure to imipenem at sub-minimum inhibitory concentration leads to altered expression of major outer membrane proteins in Acinetobacter baumannii.

AIMS: Acinetobacter baumannii is a nosocomial pathogen known to be multidrug-resistant (MDR), especially to drugs of the carbapenem class. Several factors contribute to resistance, including efflux pumps, ß-lactamases, alteration of target sites, and permeability defects. In addition, outer membrane proteins (OMPs), like porins are involved in the passage of antibiotics, and their alteration could lead to resistance development. This study aimed to explore the possible involvement of porins and OMPs in developing carbapenem resistance due to differential expression. METHODS AND RESULTS: The antibiotic-susceptible and MDR isolates of A. baumannii were first studied for differences in their transcriptional levels of OMP expression and OMP profiles. The antibiotic-susceptible isolates were further treated with imipenem, and it was found that the omp genes were differentially expressed. Six of the nine genes studied were upregulated at 1 h of exposure to imipenem. Their expression gradually decreased with time, further confirmed by their OMP profile and two-dimensional gel electrophoresis. CONCLUSIONS: This study could identify OMPs that were differentially expressed on exposure to imipenem. Hence, this study provides insights into the role of specific OMPs in antibiotic resistance in A. baumannii.

Outer Membrane Proteins and Efflux Pumps Mediated Multi-Drug Resistance in Salmonella: Rising Threat to Antimicrobial Therapy.

Despite colossal achievements in antibiotic therapy in recent decades, drug-resistant pathogens have remained a leading cause of death and economic loss globally. One such WHO-critical group pathogen is Salmonella. The extensive and inappropriate treatments for Salmonella infections have led from multi-drug resistance (MDR) to extensive drug resistance (XDR). The synergy between efflux-mediated systems and outer membrane proteins (OMPs) may favor MDR in Salmonella. Differential expression of the efflux system and OMPs (influx) and positional mutations are the factors that can be correlated to the development of drug resistance. Insights into the mechanism of influx and efflux of antibiotics can aid in developing a structurally stable molecule that can be proficient at escaping from the resistance loops in Salmonella. Understanding the strategic responsibilities and developing policies to address the surge of drug resistance at the national, regional, and global levels are the needs of the hour. In this Review, we attempt to aggregate all the available research findings and delineate the resistance mechanisms by dissecting the involvement of OMPs and efflux systems. Integrating major OMPs and the efflux system's differential expression and positional mutation in Salmonella may provide insight into developing strategic therapies for one health application.

Integrons as the potential targets for combating multidrug resistance in Enterobacteriaceae using CRISPR- Cas9 technique.

The emergence of multi-drug resistance (MDR) to pan-drug resistance (PDR) in Enterobacteriaceae has made treatment extremely challenging. Genetic mutations and horizontal gene transfer (HGT) through mobile genetic elements (MGEs) were frequently associated mechanisms of drug resistance in pathogens. However, transposons, plasmids, and integrons transfer MDR genes in bacterium via HGT much faster. Integrons are dsDNA segment that plays a crucial role in the adaptation and evolution of bacteria. They contain multiple gene cassettes that code for antibiotic resistance determinants that are expressed by a single promoter (Pc). Integrons are the cause of drug resistance in Enterobacteriaceae. Although alternatives to antibiotics such as bacteriophages, phage proteins, antimicrobial peptides, and natural compounds have been widely used to treat MDR infections, there have been limited efforts to reverse the antibiotic resistance ability of bacteria. Thus, silencing the genes harboured on MGEs achieved by Gene Editing Techniques (GETs) might prevent the spread of MDR. One such GETs, which has a simple design, good repeatability, low cost, and high efficiency, is CRISPR- Cas9 system. Thus, this review is a first of the kind that focuses on utilizing the structure of an integron to make it an ideal target for GETs like CRISPR- Cas9 systems.

Differential expression of outer membrane proteins and quinolone resistance determining region mutations can lead to ciprofloxacin resistance in Salmonella Typhi.

Multi-drug resistance in Salmonella Typhi remains a public health concern globally. This study aimed to investigate the function of quinolone resistance determining region (QRDR) of gyrA and parC in ciprofloxacin (CIP) resistant isolates and examine the differential expression of outer membrane proteins (OMPs) on exposure to sub-lethal concentrations of CIP in S. Typhi. The CIP-resistant isolates were screened for mutations in the QRDR and analyzed for bacterial growth. Furthermore, major OMPs encoding genes such as ompF, lamB, yaeT, tolC, ompS1, and phoE were examined for differential expression under the sub-lethal concentrations of CIP by real-time PCR and SDS-PAGE. Notably, our study has shown a single-point mutation in gyrA at codon 83 (Ser83-tyrosine and Ser83-phenylalanine), also the rare amino acid substitution in parC gene at codon 80 (Glu80-glycine) in CIP-resistant isolates. Additionally, CIP-resistant isolates showed moderate growth compared to susceptible isolates. Although most of the OMP-encoding genes (tolC, ompS1, and phoE) showed some degree of upregulation, a significant level of upregulation (p < 0.05) was observed only for yaeT. However, ompF and lamB genes were down-regulated compared to CIP-susceptible isolates. Whereas OMPs profiling using SDS-PAGE did not show any changes in the banding pattern. These results provide valuable information on the QRDR mutation, and the difference in the growth, and expression of OMP-encoding genes in resistant and susceptible isolates of S. Typhi. This further provides insight into the involvement of QRDR mutation and OMPs associated with CIP resistance in S. Typhi.

Non-clinical isolates of Vibrio parahaemolyticus harbouring traits of potential pathogenicity and fitness: A molecular analysis.

Here we investigated the distribution of virulence and fitness attributes V. parahaemolyticus isolated from marine environment (n = 105). We discovered ∼1% of isolates positive for tdh, 8.57% for trh, and 4.76% had tdh and trh genes. More than 50% of the isolates had pathogenicity islands specific to pandemic clones and secretion systems which are detected partially or entirely. VPaI-1 found in 59.04%; VPaI-4 in 60%; VPaI-5 in 34.28%; VPaI-2 in 99.04%; VPaI-3 in 91.42% and VPaI-6 in 99.04% isolates. Also, 34.28% of the isolates harboured T3SS2 encoding VPaI 7; T3SS1 in 98.09%; T6SS2 in 99.04% isolates and T6SS1 in 60.95% isolates. The cytotoxicity analysis showed a significant effect by causing when infected with trh+ environmental isolates. The expression of the trh, VopC, and VopA genes during infection showed a significant upregulation. This suggests the presence of virulence traits among V. parahaemolyticus that could threaten public health.

Sequence-Specific Gene Silencing of acrA in the Multi-drug Efflux System AcrAB Induces Sensitivity in Drug-Resistant Klebsiella pneumoniae.

Multi-drug efflux is one of the resistant determinants in Klebsiella pneumoniae that are encountered in a broad range of clinically relevant antimicrobial agents. An alternative method to strategically induce sensitivity in drug-resistant K. pneumoniae and improve the efficacy of the existing antibiotics is the need of the hour. Hence, an antisense RNA was designed against the acrA gene of the AcrAB-TolC efflux system in a drug-resistant isolate of K. pneumoniae obtained from a blood culture. Minimum inhibitory concentration by E test demonstrated that the antisense RNA could significantly increase the susceptibility of previously resistant K. pneumoniae toward ciprofloxacin (CIP) and co-trimoxazole. Real-time PCR determined the ability of the antisense RNA to inhibit the expression of the acrA-mRNA. The wild-type K. pneumoniae showed increased growth in the presence of CIP, while, under the same condition, the growth of the antisense RNA-treated K. pneumoniae was inhibited up till 12 h. In the presence of co-trimoxazole, delayed growth rate of the antisense RNA-treated K. pneumoniae was seen, in comparison to that of the wild-type K. pneumoniae and also a fourfold reduction was noted in the expression of the efflux gene acrA. Our results underscore the potential of the acrA antisense RNA as an alternative therapeutic against multi-drug-resistant K. pneumoniae.

Wild-type and cancer-prone zebrafish exhibit distinct gut microbial diversity and differential anti-inflammatory response upon infection.

The study investigated the gut microbial diversity and the role of gut-associated microorganisms in modulating the immune responses in normal (wild-type) and TP53M214K (cancer-prone) zebrafish. Biochemical tests, genus/species-specific PCR, and 16S rDNA sequencing were performed to characterize the bacteria isolated from the gut of wild-type (WT) and cancer-prone zebrafish. Gut microbiome analysis revealed greater diversity but reduced bacterial load in wild-type zebrafish compared with cancer-prone zebrafish, which had lesser diversity but higher bacterial load. Interestingly, the gut in cancer-prone fish showed selective colonization by opportunistic pathogens. The bacterial isolates showed resistance to antibiotics such as tetracycline, nalidixic acid, and ciprofloxacin. Gnotobiotic zebrafish embryos were established, and mono-colonization with the isolated bacteria was done to examine the expression of anti-inflammatory genes using real-time PCR. Variable expression of IL10 and IL4 was observed in germ-free (GF) wild-type embryos when mono-colonized with Staphylococcus sciuri and Vibrio cholerae. In contrast, germ-free TP53 mutant embryos showed a consistent downregulation of both the anti-inflammatory genes. Thus, a better immune response in WT embryos against S. sciuri or V. cholerae infection than in cancer-prone fish was observed, suggesting that genetic predisposition could contribute to disabling the immune system against infection.

Effect of NaCl, high iron, iron chelator and antibiotics on growth, virulence gene expression and drug susceptibility in non-typhoidal Salmonella: an in vitro fitness study.

Salmonella is one among the most versatile and resilient enteric pathogens that is known to have developed various survival strategies within the host system. The ability of the bacteria to circumvent the physiological parameters as well as dodge the antimicrobial stress environment within the host is one of the most crucial steps in establishing an infection. With an alarming rise in multi-drug resistant serovars of non-typhoidal Salmonella and lack of vaccine for combatting the infections, behaviour of the bacteria in the presence of host physiological conditions (NaCl, high and low iron) and antibiotics will help in understanding the survival strategies as well as mechanisms of resistance. Two multi-drug resistant and two sensitive serovars of Salmonella Weltevreden and Salmonella Newport isolated from poultry and seafood were used for growth kinetics and virulence gene expression study. The results obtained revealed that despite similar resistance pattern, effect of individual class of antibiotics on the growth of serovars varied. On the contrary, no significant difference was observed in growth pattern on exposure to these in vitro experimental conditions. Nevertheless, coupling these conditions with antibiotics drastically reduced the minimum inhibitory concentration (MIC) of antibiotics in resistant strains. A first of its kind study that draws attention on the significant effect of antibiotics and physiological conditions on MIC between resistant and sensitive non-typhoidal Salmonella serovars and expression of virulence genes from Salmonella pathogenicity island (SPI) 1 and 2 (invA, hilC, fliC2, sseA and ssrB).

Survival and Virulence Potential of Drug-Resistant E. coli in Simulated Gut Conditions and Antibiotic Challenge.

The gut forms a vital niche for the survival and replication of drug-resistant E. coli; however, the role of gut conditions on drug-resistant and sensitive E. coli is not clearly understood. The study aims to understand the effect of in vitro gut conditions on the spread of antibiotic resistance among E. coli and their ability to adapt to gut conditions. In this study, a multidrug-resistant (J51) and a sensitive (J254) E. coli isolate were exposed to a series of in vitro gut conditions and their growth pattern, virulence gene expression and invasion ability were studied. Further, the effect of antibiotic under in vitro gut conditions was also studied. Bile significantly affected the growth of the isolates, and the addition of iron chelator extended the lag phase of the sensitive isolate. Each in vitro gut condition had a differential effect on the expression of virulence genes in both the isolates. Further, the resistant isolate could adhere to and invade Caco2 cell lines better than the sensitive isolate. Most of the downregulated genes showed increased expression upon ciprofloxacin shock under in vitro gut conditions. The transcriptomics study revealed that exposure to bile, led to the downregulation of genes involved in different metabolic pathways. Further downregulation of metabolic pathways on ciprofloxacin shock was also observed. The downregulation of metabolic pathways could be a part of the global response played by the bacteria to adapt to harsh conditions. Reverting these fluctuated pathways could prove to be a novel strategy in combating AMR threat. Overall, bile, in high and low temperature conditions, showed a significant effect on modulating virulence gene expression on the antibiotic challenge. Thus, it is essential to consider the impact of gut conditions on gut pathogens, such as E. coli, before prescribing antimicrobial therapy during infection.

Characterisation of broad-spectrum phiKZ like jumbo phage and its utilisation in controlling multidrug-resistant Pseudomonas aeruginosa isolates.

The emergence of highly virulent multidrug-resistant P. aeruginosa has become increasingly evident among hospital-acquired infections and has raised the need for alternative therapies. Phage therapy can be one such alternative to antibiotic therapy to combat multidrug-resistant pathogenic bacteria, but this requires the availability of phages with a broad host range. In this study, isolation and molecular characterisation of P. aeruginosa specific phages were carried out. A total of 17 phages isolated showed different spectra of activity and efficiency of lysis against 82 isolates of P. aeruginosa obtained from clinical samples (n = 13), hospital effluent (n = 46) and fish processing plant effluent (n = 23). Antibiotic susceptibility test results revealed multi-drug resistance in 61 of the total 82 isolates. Three new jumbo lytic P. aeruginosa specific broad host range phages were isolated and characterised in this present study belonged to the family Myoviridae (order Caudovirales). The genetic analysis of ɸU5 revealed that phage has a genome size of 282.6 kbp with 373 putative open reading frames (ORFs), and its genetic architecture is similar to phiKZ like jumbo phages infecting P. aeruginosa. The bacteriophages isolated in this study had lytic ability against biofilm-forming and multidrug-resistant P. aeruginosa and could be candidates for further studies towards phage therapy.

Effect of sub-minimum inhibitory concentration of ceftriaxone on the expression of outer membrane proteins in Salmonella enterica serovar Typhi.

Multi-drug resistance (MDR) in Salmonella is one of the major reasons for foodborne outbreaks worldwide. Decreased susceptibility of Salmonella Typhi to first-line drugs such as ceftriaxone, ciprofloxacin, and azithromycin has raised concern. Reduced outer membrane proteins (OMPs) permeability and increased efflux pump transportation are considered to be the main reasons for the emergence of antibiotic resistance in Salmonella. The present study aimed to assess the expression of OMPs at sub-lethal concentrations of ceftriaxone in S. Typhi (Sl5037/BC, and Sl05). The S. Typhi strains were exposed to sub-MIC and half of the sub-MIC concentrations of ceftriaxone at three different time intervals (0 min, 40 min, and 180 min) and analyzed for differential expression of OMPs. Further, the expression variation of OMP encoding genes (yaeT, ompX, lamb, ompA, and ybfM) in response to ceftriaxone was evaluated using real-time PCR. The genes like lamB, ompX, and yaeT showed significant downregulation (p < 0.05) compared to the control without antibiotic exposure, whereas ybfM and ompA showed a moderate downregulation. The expression of omp genes such as lamB, ompA, ompX, ybfM, and yaeT were found to be low in the presence of ceftriaxone, followed by time and dose-dependent. The study provides insights into the possible involvement of OMPs in drug resistance of S. Typhi, which could help develop a therapeutic strategy to combat MDR isolates of S. Typhi.

'To be, or not to be'-The dilemma of 'silent' antimicrobial resistance genes in bacteria.

Antimicrobial resistance is a serious threat to public health that dramatically undermines our ability to treat bacterial infections. Microorganisms exhibit resistance to different drug classes by acquiring resistance determinants through multiple mechanisms including horizontal gene transfer. The presence of drug resistance genotypes is mostly associated with corresponding phenotypic resistance against the particular antibiotic. However, bacterial communities harbouring silent antimicrobial resistance genes-genes whose presence is not associated with a corresponding resistant phenotype do exist. Under suitable conditions, the expression pattern of such genes often revert and regain resistance and could potentially lead to therapeutic failure. We often miss the presence of silent genes, since the current experimental paradigms are focused on resistant strains. Therefore, the knowledge on the prevalence, importance and mechanism of silent antibiotic resistance genes in bacterial pathogens are very limited. Silent genes, therefore, provide an additional level of complexity in the war against drug-resistant bacteria, reminding us that not only phenotypically resistant strains but also susceptible strains should be carefully investigated. In this review, we discuss the presence of silent antimicrobial resistance genes in bacteria, their relevance and their importance in public health.

A rare occurrence of multidrug-resistant environmental Acinetobacter baumannii strains from the soil of Mangaluru, India.

Over the last decade, Acinetobacter baumannii has emerged as one of the main causes of infections acquired in the hospital setting. Outbreaks associated with this pathogen are caused mainly due to contamination and transmission in hospital territories. However, the natural habitats of A. baumannii of clinical significance still remain unclear. In this study, we highlight the isolation and identification of multidrug-resistant environmental strains of A. baumannii from the soil of Mangaluru city. All the recovered isolates were biofilm formers, and two isolates were multidrug-resistant and showed resistance to fluoroquinolone, aminoglycosides, sulfonamide, tetracycline, and carbapenems. In addition, they exhibited protease activity, and produced phospholipase C and siderophore. To the best of our knowledge, this is the first study to isolate and identify drug-resistant strains of A. baumannii from the soil.

Biofilm Formation and Associated Gene Expression in Multidrug-Resistant Klebsiella pneumoniae Isolated from Clinical Specimens.

Biofilms reduce the bacterial growth rate, inhibit antibiotic penetration, lead to the development of persister cells and facilitate genetic exchange. The biofilm-associated Klebsiella pneumoniae infections have not been well studied, and their implications in overcoming the effects of antimicrobial therapy are yet to be fully understood. Hence this study evaluated the antibiotic resistance pattern, antibiotic resistance determinants of extended-spectrum beta-lactamase (ESBL) family. Biofilm-forming ability of seventy multidrug-resistant clinical isolates of K. pneumoniae and the biofilm-associated genes of representative biofilm formers from a tertiary care hospital were also assessed. The K. pneumoniae isolated from urine exhibited resistance towards ceftazidime, nalidixic acid and meropenem. Isolates from blood were resistant to cefuroxime. Higher rates of resistance were observed towards cefuroxime, nalidixic acid, and meropenem for the isolates from the endotracheal aspirate. Extended spectrum beta-lactamase production by CLSI's disc diffusion-based confirmation test revealed all the K. pneumoniae to be as ESBL producers. Most of the isolates harboured the bla gene variants, blaSHV and blaTEM. Majority of the isolates were colistin sensitive. 97.1% of the K. pneumoniae produced biofilm. K. pneumoniae isolated from pus and blood produced fully established biofilms. Strong biofilm formers were sensitive to co-trimoxazole and ciprofloxacin. Moderate biofilm formers exhibited sensitivity towards meropenem and imipenem. Expression of the fimH gene was increased, while mrkD showed reduced expression among the strong biofilm formers. Moderate biofilm formers showed variable expression of the genes associated with the biofilm formation. The weak and non-biofilm formers showed reduced expression of both the fimbrial genes. Multidrug-resistant isolates produced ESBLs and formed well-established biofilms.

Effect of ciprofloxacin and in vitro gut conditions on biofilm of Escherichia coli isolated from clinical and environmental sources.

AIM: This study aimed at characterizing the biofilm-forming ability of drug-resistant and sensitive Escherichia coli under in vitro gut conditions and in the presence of ciprofloxacin. METHODS AND RESULTS: 153 E. coli isolates comprising 80 from clinical and 73 from environment source were studied for their ability to form biofilm under control and in vitro simulated gut conditions. The integrity of preformed biofilm on exposure to ciprofloxacin was assessed. Expression of biofilm-associated genes was analysed using qPCR. A high degree of resistance was observed in clinical isolates with a concomitant prevalence of blaTEM . Bile, pH and low temperature enabled the E. coli biofilm to resist the effect of ciprofloxacin. Clinical isolates of E. coli formed strong biofilms in in vitro gut conditions following exposure to high concentration of ciprofloxacin. The expression of biofilm genes varied between different gut conditions viz., presence of bile, pH and low temperature, included in this study. CONCLUSIONS: This study demonstrates the importance of papC and csgA for maintaining the biofilm integrity upon antibiotic exposure. Escherichia coli form biofilm as a survival strategy to adapt to the conditions in their environment irrespective of their drug resistance status. SIGNIFICANCE AND IMPACT OF THE STUDY: The study provides an understanding of the effect of different parameters of the gut conditions during infection and the effect of antibiotic on survival and biofilm-forming ability of clinical and environmental E. coli isolates. It further suggests that bacteria resort to biofilm formation as one of the mechanisms to adjust to alterations in gut conditions and once the biofilm is formed, it requires high concentration of ciprofloxacin to eradicate it.

Comparative analysis of different methods used for molecular characterization of Burkholderia cepacia complex isolated from noncystic fibrosis conditions.

PURPOSE: Burkholderia is a Gram-negative opportunistic bacterium capable of causing severe nosocomial infections. The aim of this study was to characterize Burkholderia cepacia complex and to compare different molecular methods used in its characterization. METHODS: In this study, 45 isolates of Burkholderia cepacia complex (Bcc) isolated from clinical cases were subjected to RAPD (Random amplified polymorphic DNA), recA-RFLP (Restriction fragment length polymorphism), 16SrDNA-RFLP, whole-cell protein analysis, recA DNA sequencing and biofilm assay. RESULTS: Of the 45 isolates tested, 97.7% were sensitive to ceftazidime, 82.2% were sensitive to Cotrimoxazole, 73.3% were sensitive to meropenem, 55.5% were sensitive to minocycline and 42.2% were sensitive to levofloxacin. Majority of the isolates harbored all the tested virulence genes except bpeA and cblA. The RAPD generated 11 groups (R1-R11), recA-RFLP 10 groups (A1-A10), 16SrRNA-RFLP 5 groups (S1-S5) and SDS-PAGE (Sodium Dodecyl Sulphate-Polyacrylamide gel electrophoresis) whole cell protein analysis revealed 12 groups (C1-C12). recA sequencing revealed that most of the isolates belonging to the genomovar III Burkholderia cenocepacia. Though all the methods are found to be efficient in differentiating Burkholderia spp., recA-RFLP was highly discriminatory at 96% similarity value. The study also identified a new strain Burkholderia pseudomultivorans for the first time in the country. Further, recA sequencing could identify the strains to species level. Majority of the multidrug-resistant strains also showed moderate to strong biofilm-forming ability, which further contributes to the virulence characteristics of the pathogens. CONCLUSIONS: The study highlights the importance of combination of molecular methods to characterize Burkholderia cepacia complex. Molecular typing of these human pathogens yields important information for the clinicians in order to initiate the most appropriate therapy in the case of severe infections and to implement preventive measures for the effective control of transmission of Burkholderia spp.

Molecular investigation of the dengue outbreak in Karnataka, South India, reveals co-circulation of all four dengue virus serotypes.

The growing incidence of dengue outbreaks in the state of Karnataka prompted us to study the circulating dengue virus (DENV) and their proportion among the suspected cases of dengue patients during the disease outbreak at Mysuru district of Southern India. The presence of the DENV in a patient's serum sample was identified by RT-PCR using previously published primer pairs targeting CprM gene. DENV serotyping was carried out by semi-nested multiplex PCR using serotype-specific primers and nucleotide sequencing. Three hundred fifty-five samples of serum from suspected dengue cases were collected, and 203 samples (57.18%) were found positives. In 2016, DENV-4 (97.87%) was found to be the most dominant DENV serotype either alone or as co-infection, followed by DENV-2 (8.51%) and DENV-3 (4.25%). In 47 positive cases, co-infection with more than one serotype was detected in 4 cases (8.51%). The analysis of the dengue cases in 2017, DENV-4 was dominating serotype (33.97%), followed by the emergence of DENV-2 (32.05%), DENV-3 (25.64%), and DENV-1 (25.00%). Our study also reports the circulation of all four DENV serotypes in the Mysuru district of Southern India, with concurrent infections rate of 16.66% in 2017. The present study provides information regarding the genetic variation among the circulating DENV serotype in an Indian state of Karnataka. The need for the studying genetic diversity of DENV will be useful during the continuous monitoring for disease burden as well as the development of appropriate prophylactic measures to control the spread of dengue infection.

Phenotypic characterization of auxotrophic mutant of nontyphoidal Salmonella and determination of its cytotoxicity, tumor inhibiting cytokine gene expression in cell line models.

An auxotrophic mutant of nontyphoidal Salmonella (NTS) strain (Salmonella Oslo) was phenotypically characterized in this study. The characterization was based on phenotype, morphology, motility, biofilm forming ability, growth kinetics, etc. The phenotypic results from the above experiments determined that the mutant showed variation in phenotypic characters from that of wild-type strain. Subsequently, mutant and wild-type NTS were subjected to epithelial cell invasion and intracellular replication assays. The real-time PCR analysis was also performed to analyse expression of tumor inhibiting cytokine genes and virulence genes post-bacterial infection in cell lines. The mutant showed highest invasion potential than wild-type NTS whereas the replication of mutant was slower in both the cell lines. Similar to the wild-type strain, the mutant also retained the cytotoxic potential when analysed in vitro. Furthermore, the expression of proinflammatory cytokine genes such as TNF-α and IL-1ß was upsurged with the downregulation of anti-inflammatory cytokine genes like TGF-ß, IL-6 and IL-10 post-infection of the mutant strain in cell lines. In addition, virulence genes of Salmonella pathogenicity island one and two of mutant were downregulated in vitro except invA in HeLa cell line. Therefore, the auxotrophic mutant showed positive attributes of a potential antitumor agent in terms of expressing tumor inhibiting cytokine genes when assessed in vitro. Though the study did not check the tumor inhibitory effect of NTS strain directly, findings of the study emphasizes on the development of a novel strain of NTS with less virulence and more immunogenic traits to inhibit tumor cells.

Detection technologies and recent developments in the diagnosis of COVID-19 infection.

COVID-19 is a disease caused by SARS-CoV-2 capable of causing mild to severe infections in humans. Since its first appearance in China in December 2019, the pandemic has spread rapidly throughout the world. Despite considerable efforts made to contain the disease, the virus has continued its prevalence in many countries with varying degrees of clinical manifestations. To contain this pandemic, collaborative approach involving accurate diagnosis, epidemiology, surveillance, and prophylaxis is essential. However, proper diagnosis using rapid technologies plays a crucial role. With increasing incidence of COVID-19 cases, the accurate and early detection of the SARS-CoV-2 is need of the hour for effective prevention and management of COVID-19 cases as well as to curb its spread. RT-qPCR assay is considered to be the gold standard for the early detection of virus, but this protocol has limited application to use as bedside test because of its technical complexity. To address these challenges, several POC assays have been developed to facilitate the COVID-19 diagnosis outside the centralized testing laboratories as well to accelerate the clinical decision making with a least turnaround time. Hence, in this report, we review different nucleic acid-based and serological techniques available for the diagnosis and effective prevention of COVID-19. KEY POINTS : • Provides comprehensive information on the different diagnostic tools available for COVID-19 • Nucleic acid based tests or antigen detection tests are used for diagnostic purpose • Accurate diagnosis is essential for the efficient management of COVID-19.

Burkholderia vietnamiensis causing a non-lactational breast abscess in a non-cystic fibrosis patient in Tamil Nadu, India.

Burkholderia cepacia complex is a Gram-negative opportunistic pathogen usually found in people with an immunocompromised condition such as cystic fibrosis (CF). In a tropical country like India, this organism has been associated with a number of hospital-acquired infections including sepsis. We present here a report of a case of Burkholderia vietnamiensis causing a non-lactational breast abscess in a non-CF patient. The pathogen was identified as B. cepacia using Vitek system and matrix-assisted laser desorption ionisation-time of flight. This was confirmed by polymerase chain reaction (PCR) using recA genus-specific gene and sequencing of the PCR amplicons. recA-restriction fragment length polymorphism and recA gene sequencing revealed that the isolate is B. vietnamiensis. This is the first description of B. vietnamiensis isolated from a clinical case from India.