Identification of circulating miRNA biomarkers in Leptospirosis for early detection: A Promising Diagnostic Approach.

Leptospirosis is a zoonotic disease of global significance, contributing to morbidity and mortality worldwide. It is endemic to tropical regions, with outbreaks during monsoons. The disease manifestations are similar to that of other febrile illness such as dengue, malaria hence often misdiagnosed and underreported. The zoonoses if undetected, progresses to cause severe life-threatening complications also known as Weil's disease. Routine diagnostic tests are based on the detection of antibodies in patient serum and are not accurate during the initial phase of the infection. Therefore, it is necessary to detect novel biomarkers that can be used in early detection of leptospirosis. Circulating miRNAs are known to be promising biomarkers for various diseases including cancer, tuberculosis, influenza; hence in this study the potential of miRNAs as biomarkers for leptospirosis was evaluated. A total of 30 leptospirosis cases were screened for the differential expression of 10 miRNA by RT-qPCR assay. The differential expression was calculated by relative quantification using healthy individuals as controls. Among the 10 miRNA,3 miRNA, miR-28-5p, miR-302c-3p and miR-302a-3p were reported to exhibit a significant trend of upregulation. Further their role in immune pathways and biological processes was investigated by KEGG analysis and Gene Ontology. The 3 miRNAs were observed to target various immune response pathways, thus confirming their role in host immune response. Based on the results obtained in this study, miR-28-5p, miR-302c-3p and miR-302a-3p can be considered as potential biomarkers for the detection of leptospirosis.

Oral administration of recombinant outer membrane protein A-based nanovaccine affords protection against Aeromonas hydrophila in zebrafish.

Aeromonas hydrophila, an opportunistic warm water pathogen, has always been a threat to aquaculture, leading to substantial economic losses. Vaccination of the cultured fish would effectively prevent Aeromoniasis, and recent advancements in nanotechnology show promise for efficacious vaccines. Oral delivery would be the most practical and convenient method of vaccine delivery in a grow-out pond. This study studied the immunogenicity and protective efficacy of a nanoparticle-loaded outer membrane protein A from A. hydrophila in the zebrafish model. The protein was over-expressed, purified, and encapsulated using poly lactic-co-glycolic acid (PLGA) nanoparticles via the double emulsion method. The PLGA nanoparticles loaded with recombinant OmpA (rOmpA) exhibited a size of 295 ± 15.1 nm, an encapsulation efficiency of 72.52%, and a polydispersity index of 0.292 ± 0.07. Scanning electron microscopy confirmed the spherical and isolated nature of the PLGA-rOmpA nanoparticles. The protective efficacy in A. hydrophila-infected zebrafish after oral administration of the nanovaccine resulted in relative percentage survival of 77.7. Gene expression studies showed significant upregulation of immune genes in the vaccinated fish. The results demonstrate the usefulness of oral administration of nanovaccine-loaded rOmpA as a potential vaccine since it induced a robust immune response and conferred adequate protection against A. hydrophila in zebrafish, Danio rerio.

Identification of Dominant Leptospira Serogroups among Leptospirosis Cases and Their Clinical Outcomes: A Prospective Hospital-Based Study in Mangaluru, India.

Leptospirosis is a reemerging zoonotic disease of worldwide significance, endemic to the southern region of India, with clinical manifestations similar to other febrile illnesses; hence, it is often misdiagnosed and underreported. Inadequate information about the disease burden and the regional circulating serogroups contributes to its neglected disease status. This study aimed to identify the infecting Leptospira serogroup in the coastal region of Mangaluru and study the clinical symptoms and outcome among leptospirosis patients. Serum samples were collected from 30 patients with confirmed leptospirosis admitted to a tertiary care center in Mangaluru and screened by microscopic agglutination test (MAT) for the infecting serogroup. The clinical profile of these cases was reviewed, and data regarding epidemiological factors such as age, sex, complications, and mortality were recorded. The MAT identified a higher occurrence of serogroup Bataviae (n = 7, 43.75%) and serogroup Australis (n = 5, 31.25%) compared with other serogroups screened in this study population. Patients were aged 16 to 65 years, with a predominance of males. The clinical presentation of leptospirosis ranged from a mild febrile illness to multiorgan failure. Fever (n = 29, 96%) was the common clinical presentation, followed by myalgia, nausea, and abdominal pain. Acute kidney injury, acute respiratory distress syndrome, and multiple organ dysfunction syndrome were the common complications observed. Determining the circulating serogroup is necessary to understand the epidemiology and diversity of Leptospira serogroups among animals and humans to strategize appropriate preventive measures.

Polylactic-Co-glycolic Acid Polymer-Based Nano-Encapsulation Using Recombinant Maltoporin of Aeromonas hydrophila as Potential Vaccine Candidate.

Aquaculture production has been incurring economic losses due to infectious diseases by opportunistic pathogens like Aeromonas hydrophila, a bacterial agent that commonly affects warm water aquacultured fish. Developing an effective vaccine with an appropriate delivery system can elicit an immune response that would be a useful disease management strategy through prevention. The most practical method of administration would be the oral delivery of vaccine developed through nano-biotechnology. In this study, the gene encoding an outer membrane protein, maltoporin, of A. hydrophila, was identified, sequenced, and studied using bioinformatics tools to examine its potential as a vaccine candidate. Using a double emulsion method, the molecule was cloned, over-expressed, and encapsulated in a biodegradable polymer polylactic-co-glycolic acid (PLGA). The immunogenicity of maltoporin was identified through in silico analysis and thus taken up for nanovaccine preparation. The encapsulation efficiency of maltoporin was 63%, with an in vitro release of 55% protein in 48 h. The particle size and morphology of the encapsulated protein exhibited properties that could induce stability and function as an effective carrier system to deliver the antigen to the site and trigger immune response. Results show promise that the PLGA-mediated delivery system could be a potential carrier in developing a fish vaccine via oral administration. They provide insight for developing nanovaccine, since sustained in vitro release and biocompatibility were observed. There is further scope to study the immune response and examine the protective immunity induced by the nanoparticle-encapsulated maltoporin by oral delivery to fish.

In vitro determination of probiotic efficacy of Bacillus subtilis TLDK301120C24 isolated from tilapia against warm water fish pathogens and in vivo validation using gnotobiotic zebrafish model.

Eco-friendly alternatives such as probiotics are needed to prevent economically relevant infectious diseases for a successful disease-free harvest in aquaculture. The use of antibiotics has been the favored practice, but its empirical and indiscriminate use has led to antibiotic resistance in the aquatic environment and residues in the food fish. With this rationale, a probiotic was isolated from tilapia, a commercially important cultured fish worldwide. The characteristics of the probiotic were checked against common bacterial pathogens affecting aquaculture. In vitro tests demonstrated the inhibitory effects of the isolated probiotic on the growth of Aeromonas hydrophila, Edwardsiella tarda, Vibrio anguillarum, and V. alginolyticus. The candidate probiotic, referred to as TLDK301120C24, was identified as Bacillus subtilis by a battery of biochemical tests and genotypic confirmation by 16S rDNA sequencing. The in vitro results revealed the ability of the probiotic to withstand the gut conditions that included pH range of 3-9, salt concentration of 0.5-6%, and bile salt concentration of up to 6%. The isolate could hydrolyze starch (12-14 mm clearance zone), protein (20-22 mm clearance zone), and cellulose (22-24 mm clearance zone). Further, the inhibitory ability of the probiotic against aquatic pathogens was determined in vivo using gnotobiotic zebrafish by employing a novel approach that involved tagging the probiotic with a red fluorescent protein and the pathogens with a green fluorescent protein, respectively. The colonizing ability of probiotics and its inhibitory effects against the pathogens were evaluated by fluorescence microscopy, PCR, and estimation of viable counts in LBA + Amp plates. Finally, the competitive inhibition and exclusion of fish pathogens A. hydrophila and E. tarda by B. subtilis was confirmed semi-quantitatively, through challenge experiments. This study shows the potential of B. subtilis as a probiotic and its excellent ability to inhibit major fish pathogens in vivo and in vitro. It also shows promise as a potent substitute for antibiotics.

Nanovaccines to Combat Aeromonas hydrophila Infections in Warm-Water Aquaculture: Opportunities and Challenges.

The application of nanotechnology in aquaculture for developing efficient vaccines has shown great potential in recent years. Nanovaccination, which involves encapsulating antigens of fish pathogens in various polymeric materials and nanoparticles, can afford protection to the antigens and a sustained release of the molecule. Oral administration of nanoparticles would be a convenient and cost-effective method for delivering vaccines in aquaculture while eliminating the need for stressful, labour-intensive injectables. The small size of nanoparticles allows them to overcome the degradative digestive enzymes and help deliver antigens to the target site of the fish more effectively. This targeted-delivery approach would help trigger cellular and humoral immune responses more efficiently, thereby enhancing the protective efficacy of vaccines. This is particularly relevant for combating diseases caused by pathogens like Aeromonas hydrophila, a major fish pathogen responsible for significant morbidity and mortality in the aquaculture sector. While the use of nanoparticle-based vaccines in aquaculture has shown promise, concerns exist about the potential toxicity associated with certain types of nanoparticles. Some nanoparticles have been found to exhibit varying degrees of toxicity, and their safety profiles need to be thoroughly assessed before widespread application. The introduction of nanovaccines has opened new vistas for improving aquaculture healthcare, but must be evaluated for potential toxicity before aquaculture applications. Details of nanovaccines and their mode of action, with a focus on protecting fish from infections and outbreaks caused by the ubiquitous opportunistic pathogen A. hydrophila, are reviewed here.

Metagenomic Analysis to Uncover the Subgingival and Atherosclerotic Plaque Microbiota in Patients with Coronary Artery Disease.

The role of periodontal pathogens in the initiation and progression of atherosclerosis has been extensively researched, yet a precise causal mechanism has not been established. The subgingival microbiota may be a source of dissemination and may contribute to the development of atherosclerosis; hence this study attempted to characterize and compare the subgingival and atherosclerotic plaques. Plaque samples were subjected to 16S rRNA-based metagenomics to study microbiota associated with subgingival and atherosclerotic plaques collected from patients with coronary artery disease. The PCoA analysis showed that the microbiomes of subgingival plaques were highly scattered and showed a diverse microbial composition, unlike the atherosclerotic plaques that did not show evident variability in the microbial composition and formed a close distinct group. The abundance of various genera in the subgingival plaques revealed Fusobacterium (11%), Acinetobacter (13%), Veillonella (9%), and Prevotella (11%) among the top ten genera. The atherosclerotic plaques contained Acinetobacter (39%), Chryseobacterium (9%), Rhizobium (5%), and Staphylococcus (4%). All the patients examined in this study had either generalized or localized periodontitis with varying degrees of severity. The community microbiota analysis revealed that 22 bacterial genera were shared between two different plaques, with Acinetobacter being dominant. Based on the Human Oral Microbiome Database, 55% of the shared microbiota in this study have been listed as periodontal microbiota, with some of them found in increased proportions in patients with periodontitis suggesting the translocation of bacteria from the periodontal pockets into the circulation. This study provides valuable insights into the possible relationship between periodontal pathogens and atherosclerotic cardiovascular disease.

Genome analysis of clinical genotype Vibrio vulnificus isolated from seafood in Mangaluru Coast, India provides insights into its pathogenicity.

Vibrio vulnificus an opportunistic human pathogen native to marine/estuarine environment, is one of the leading causes of death due to seafood consumption and exposure of wounds to seawater worldwide. The present study involves the whole genome sequence analysis of an environmental strain of V. vulnificus (clinical genotype) isolated from seafood along the Mangaluru coast of India. The sequenced genome data was subjected to in-silico analysis of phylogeny, virulence genes, antimicrobial resistance determinants, and secretary proteins using suitable bioinformatics tools. The sequenced isolate had an overall genome length of 4.8 Mb and GC content of 46% with 4400 coding DNA sequences. The sequenced strain belongs to a new sequence type (Multilocus sequence typing) and was also found to branch with a phylogenetic lineage that groups the most infectious strains of V. vulnificus. The seafood isolate had complete genes involved in conferring serum resistance yet showed limited serum resistance. The study identified several genes against the antibiotics that are commonly used in their treatment, highlighting the need for alternative treatments. Also, the secretory protein analysis revealed genes associated with major pathways like ABC transporters, two-component systems, quorum sensing, biofilm formation, cationic antimicrobial peptide (CAMP) resistance, and others that play a critical role in the pathogenesis of the V. vulnificus. To the best of our knowledge, this is the first report of a detailed analysis of the genomic information of a V. vulnificus isolated from the Indian subcontinent and provides evidence that raises public health concerns about the safety of seafood.

Evaluation of Probiotic Efficacy of Bacillus subtilis RODK28110C3 Against Pathogenic Aeromonas hydrophila and Edwardsiella tarda Using In Vitro Studies and In Vivo Gnotobiotic Zebrafish Gut Model System.

The indiscriminate use of antibiotics in aquaculture has led to the emergence of resistance; hence, eco-friendly, host-specific alternatives to mitigate bacterial infections have become imminent. In this study, bacteria that could possibly serve as probiotics were isolated and evaluated for their efficacy with in vitro experiments and in vivo zebrafish gut model. One isolate from each of the 23 rohu fish (Labeo rohita) was shortlisted after preliminary screening of several isolates and tested for their ability to inhibit two important warm water bacterial fish pathogens, Aeromonas hydrophila, and Edwardsiella tarda. An isolate (RODK28110C3) that showed broad-spectrum inhibitory activity against a battery of different isolates of the two fish pathogens included in this study and maintained in our repository was selected for further characterization. The culture was identified phenotypically as Bacillus subtilis and confirmed by 16S rDNA sequencing. The isolate was able to hydrolyze fish feed constituents that include starch, protein, and cellulose. Further in vitro tests ensured that the potential isolate with probiotic attributes could tolerate different gut conditions, which included a range of pH, salinity, and varying concentrations of bile salt. Exposure of 4 days post fertilization zebrafish embryos to the RFP-tagged isolate confirmed the colonization of B. subtilis in the gut of the zebrafish embryo, which is an important attribute of a probiotic. The isolate was able to inhibit both A. hydrophila and E. tarda in gnotobiotic zebrafish embryo in triplicate. The study demonstrates the probiotic characteristics of the B. subtilis isolated from L. rohita and its ability to inhibit A. hydrophila and E. tarda using in vitro conditions and in the zebrafish gut and could serve as an effective alternative to antibiotics in aquaculture.

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.

An insight into clinical and laboratory detections for screening and diagnosis of cervical cancer.

INTRODUCTION: Cervical cancer is the most frequent malignancy among women caused by an unresolved long-term infection with distinct human papillomavirus (HPV) genotypes. It is the fourth most common form of cancer among women worldwide. The two oncogenic genotypes, HPV 16 and 18, are responsible for >70% of all cervical cancers worldwide. Cervical cancer is one of the most successfully preventable and treatable forms of cancer if detected early. AREAS COVERED: In this review article, we have summarizedsummarised the different approaches used in clinical diagnosis and research laboratories to detect HPV-related changes associated with cervical cancer for a better understanding of the advantages and limitations of these tests. EXPERT OPINION: Despite the well-known screening strategies for cervical cancer, developing nations lack effective implementation due to various factors. With the current rate of cervical cancer cases, precise and timely identification of HPV can significantly impact the prevention and efficient management of cervical cancer. Cervical cancer is the most common gynecological cancer in developing countries. The primary screening test with cytology and molecular testing of HPV is important for preventing cervical cancer. To address these issues, several point-of-care assays have been developed to facilitate rapid screening of HPV with the least turnaround time.

Evaluation of reverse transcriptase-polymerase spiral reaction assay for rapid and sensitive detection of severe acute respiratory syndrome coronavirus 2.

BACKGROUND AND AIM: Existing real-time reverse transcriptase PCR (RT-qPCR) has certain limitations for the point-of-care detection of severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) since it requires sophisticated instruments, reagents and skilled laboratory personnel. In this study, we evaluated an assay termed the reverse transcriptase-polymerase spiral reaction (RT-PSR) for rapid and visual detection of SARS-CoV-2. METHODS: The RT-PSR assay was optimized using RdRp gene and evaluated for the detection of SARS-CoV-2. The time of 60min and a temperature of 63°C was optimized for targeting the RNA-dependent RNA polymerase gene of SARS-CoV-2. The sensitivity of the assay was evaluated by diluting the in-vitro transcribed RNA, which amplifies as low as ten copies. RESULTS: The specific primers designed for this assay showed 100% specificity and did not react when tested with other lung infection-causing viruses and bacteria. The optimized assay was validated with 190 clinical samples in two phases, using automated RTPCR based TrueNat test, and the results were comparable. CONCLUSIONS: The RT-PSR assay can be considered for rapid and sensitive detection of SARS-CoV-2, particularly in resource-limited settings. To our knowledge, there is as yet no RT-PSR-based kit developed for SARS-CoV-2.

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.

Aeromonas species isolated from aquatic organisms, insects, chicken, and humans in India show similar antimicrobial resistance profiles.

Aeromonas species are Gram-negative bacteria that infect various living organisms and are ubiquitously found in different aquatic environments. In this study, we used whole genome sequencing (WGS) to identify and compare the antimicrobial resistance (AMR) genes, integrons, transposases and plasmids found in Aeromonas hydrophila, Aeromonas caviae and Aeromonas veronii isolated from Indian major carp (Catla catla), Indian carp (Labeo rohita), catfish (Clarias batrachus) and Nile tilapia (Oreochromis niloticus) sampled in India. To gain a wider comparison, we included 11 whole genome sequences of Aeromonas spp. from different host species in India deposited in the National Center for Biotechnology Information (NCBI). Our findings show that all 15 Aeromonas sequences examined had multiple AMR genes of which the Ambler classes B, C and D ß-lactamase genes were the most dominant. The high similarity of AMR genes in the Aeromonas sequences obtained from different host species point to interspecies transmission of AMR genes. Our findings also show that all Aeromonas sequences examined encoded several multidrug efflux-pump proteins. As for genes linked to mobile genetic elements (MBE), only the class I integrase was detected from two fish isolates, while all transposases detected belonged to the insertion sequence (IS) family. Only seven of the 15 Aeromonas sequences examined had plasmids and none of the plasmids encoded AMR genes. In summary, our findings show that Aeromonas spp. isolated from different host species in India carry multiple AMR genes. Thus, we advocate that the control of AMR caused by Aeromonas spp. in India should be based on a One Health approach.

Genome analysis of clinical isolate of Campylobacter fetus subspecies fetus MMM01 from India reveals genetic determinants of pathogenesis and adaptation.

In this study we report the whole genome sequencing (WGS) based analysis of blood-borne Campylobacter fetus subsp. fetus MMM01 isolated from a diabetic patient to obtain deeper insights in to the virulence and host adaptability. The sequenced genome of C. fetus subsp. fetus MMM01 along with reference genomes retrieved from NCBI was subjected to various in-silico analysis including JSpecies, MLST server, PATRIC server, VFanalyzer, CARD, PHASTER to understand their phylogenetic relation, virulence and antimicrobial resistance profile. The genome had a size of 1,788,790 bp, with a GC content of 33.09%, nearly identical to the reference strain C. fetus subsp. fetus 82-40. The MLST based phylogenetic tree constructed revealed the polyphyletic branching and MMM01 (ST25) was found to be closely related to ST11, both belong to the sap-A serotype which are more common in human infections. VFanalyzer identified 88 protein-coding genes coding for several virulence factors including Campylobacter adhesion to fibronectin, flagellar apparatus, cytolethal distending toxin operons and Campylobacter invasion antigen proteins which enhance the virulence of bacteria along with resistance genes against antibiotics including fluoroquinolone, chloramphenicol, tetracycline, and aminoglycoside in MMM01, which points to enhanced survival and pathogenicity of this zoonotic pathogen. It was interesting to find that MMM01 lacked FGI-II island found in most of the clinical isolates, which encoded CRISPR Cas and prophage II regions. More details about the complexity and evolution of this zoonotic pathogen could be learned from future studies that concentrate on comparative genome analysis using larger genome datasets.

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.

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).

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.

Current methods for the diagnosis of leptospirosis: Issues and challenges.

Leptospirosis is a re-emerging anthropo-zoonotic infection of worldwide significance caused by the pathogenic spirochete (Leptospira interrogans) of the genus Leptospira, predominant in tropical/temperate regions and endemic to areas receiving heavy rainfall and flooding. Clinical presentation is similar to that of other febrile illnesses exhibiting mild symptoms which are often self-limiting. Hence, Leptospirosis is often mis-diagnosed and remains untreated progressing to Weil's Disease which is fatal. As only 30% of cases are diagnosed in endemic countries, Leptospirosis remains as a neglected zoonotic disease of tropical regions, due to poor diagnostic facilities and mild, asymptomatic disease manifestations which are often neglected. As this zoonosis is reported to cause periodical outbreaks, it is a major public health concern. Although diagnostic facilities are available, they are not accessible in technology-limited settings and are limited to certain hospitals and reference laboratories. This review is about the various methods used for the detection of Leptospirosis and their significance. It highlights the need for an appropriate diagnostic test for the rapid detection of leptospirosis in order to initiate immediate antibiotic therapy.

Association of exopolysaccharide genes in biofilm developing antibiotic-resistant Pseudomonas aeruginosa from hospital wastewater.

The study aimed to examine the relationship between antibiotic resistance, biofilm formation and genes responsible for biofilm formation. Sixty-six Pseudomonas aeruginosa isolates were obtained from hospital wastewater and analyzed for their antibiotic resistance. Biofilm production among the isolates was tested by indirect quantification method crystal violet assay. Biofilm-associated genes among these isolates psl, alg, and pel were also checked. The maximum resistance was observed for ampicillins (88.24%) followed by nalidixic (83.82%), and nitrofurantoin (64.71%), respectively. Biofilm phenotypes are distributed in the following categories: high 39.39% (n = 26); moderate 57.57% (n = 38), and weak 3.0% (n = 2). Among the total isolates, biofilm-associated genes were detected in 84.84% (n = 56) of isolates and the remaining isolates 15.15% (n = 10) did not harbor any genes. In this study, pslB was the most predominant gene observed (71.21%, n = 47) followed by pslA (57.57%, n = 38), pelA (45.45%, n = 30), algD (43.93%, n = 29), and pelD (27.27%, n = 18), respectively. The present study reveals that the majority of the isolates are multidrug resistant being moderate and high biofilm formers. The study implies that biofilm acts as a machinery for bacteria to survive in the hospital effluent which is an antibiotic stress environment.