Antimicrobial resistance heterogeneity among multidrug-resistant Gram-negative pathogens: Phenotypic, genotypic, and proteomic analysis.

Microbes evolve rapidly by modifying their genomes through mutations or through the horizontal acquisition of mobile genetic elements (MGEs) linked with fitness traits such as antimicrobial resistance (AMR), virulence, and metabolic functions. We conducted a multicentric study in India and collected different clinical samples for decoding the genome sequences of bacterial pathogens associated with sepsis, urinary tract infections, and respiratory infections to understand the functional potency associated with AMR and its dynamics. Genomic analysis identified several acquired AMR genes (ARGs) that have a pathogen-specific signature. We observed that blaCTX-M-15, blaCMY-42, blaNDM-5, and aadA(2) were prevalent in Escherichia coli, and blaTEM-1B, blaOXA-232, blaNDM-1, rmtB, and rmtC were dominant in Klebsiella pneumoniae. In contrast, Pseudomonas aeruginosa and Acinetobacter baumannii harbored blaVEB, blaVIM-2, aph(3'), strA/B, blaOXA-23, aph(3') variants, and amrA, respectively. Regardless of the type of ARG, the MGEs linked with ARGs were also pathogen-specific. The sequence type of these pathogens was identified as high-risk international clones, with only a few lineages being predominant and region-specific. Whole-cell proteome analysis of extensively drug-resistant K. pneumoniae, A. baumannii, E. coli, and P. aeruginosa strains revealed differential abundances of resistance-associated proteins in the presence and absence of different classes of antibiotics. The pathogen-specific resistance signatures and differential abundance of AMR-associated proteins identified in this study should add value to AMR diagnostics and the choice of appropriate drug combinations for successful antimicrobial therapy.

Risk factors and outcome associated with the acquisition of MDR linezolid-resistant Enterococcus faecium: a report from tertiary care centre.

OBJECTIVE: The aim of the study was to determine the resistance profile of linezolid-resistant Enterococcus faecium (LREfm) and to investigate risk factors and outcomes associated with LREfm infections. MATERIAL AND METHODS: A prospective case-control study was undertaken (2019 to 2022) and included 202 patients with LREfm infections (cases) and 200 controls with LSEfm infections. Clinical data was prospectively collected and analysed for risk factors and outcomes. Antimicrobial susceptibility was performed, and resistance profile was studied using WHOnet. RESULTS: Risk factors associated with LREfm infection were site of infection UTI (OR 5.87, 95% CI 2.59-13.29, p ≤ 0.001), prior use of carbapenem (OR 2.85 95% CI 1.62-5.02, p ≤ 0.001) and linezolid (OR 10.13, 95% CI 4.13-24.82, p ≤ 0.001), use of central line (OR 5.54, 95% CI 2.35-13.09, p ≤ 0.001), urinary catheter (OR 0.29, 95% CI 0.12-0.70, p ≤ 0.001) and ventilation (OR 14.87, 95% CI 7.86-28.11, p ≤ 0.007). The hospital stay 8-14 days (< 0.001) prior to infection and the mortality rate (p = 0.003) were also significantly high among patients with LREfm infections. Linezolid and vancomycin resistance coexisted; further, MDR, XDR and PDR phenotypes were significantly higher among LREfm. CONCLUSION: This study provided insight into epidemiology of MDR LREfm in a setting where linezolid use is high. The main drivers of infections with LREfm are multiple, including use of carbapenems and linezolid. Invasive procedures and increased hospital stay facilitate spread through breach in infection control practises. As therapeutic options are limited, ongoing surveillance of LREfm and VRE is critical to guide appropriate use of linezolid and infection control policies.

Varicella zoster virus outbreak in a long-term care unit of a tertiary care hospital in northern India.

Nosocomial outbreak of varicella zoster virus (VZV) has been reported when susceptible individuals encounter a case of chicken pox or shingles. A suspected VZV outbreak was investigated in a 50-bedded in-patient facility of Physical Medicine and Rehabilitation in a tertiary care multispecialty hospital. A 30-year-old female patient admitted with Pott's spine was clinically diagnosed with chicken pox on 31 December 2022. The following week, four more cases were identified in the same ward. All cases were diagnosed as laboratory-confirmed varicella zoster infection by PCR. Primary case was a housekeeping staff who was clinically diagnosed with chicken pox 3 weeks prior (9 December 2022). He returned to work on eighth day of infection (17 December 2022) after apparent clinical recovery but before the lesions had crusted over. Thirty-one HCWs were identified as contacts a and three had no evidence of immunity. Two of these susceptible HCWs had onset of chickenpox shortly after first dose of VZV vaccination was inoculated. All cases recovered after treatment with no reported complications. VZV infection is highly contagious in healthcare settings with susceptible populations. Prompt identification of cases and implementation of infection prevention and control measures like patient isolation and vaccination are essential for the containment of outbreaks.

Evaluation of risk factors associated with hard-to-heal wound infection in a tertiary care hospital.

OBJECTIVE: The incidence of hard-to-heal wound infection, especially as a result of multidrug-resistant Gram-negative organisms, has increased in recent years. The reason for the increase is multifactorial and the ability of these pathogenic isolates to form biofilms is one of the important risk factors in wound infection. This study aimed to evaluate the risk factors associated with such cases. METHOD: This prospective analytical study, conducted over a period of two months, included pus or tissue samples from hospital inpatients with Gram-negative hard-to-heal wound infection. The samples were processed with conventional microbiological techniques. Patient demographic details and the presence of various risk factors were recorded. Biofilm production was detected by tissue culture plate method in the laboratory. The data were analysed using SPSS version 21 (IBM Ltd., US). RESULTS: The experimental cohort comprised 200 patients. Klebsiella spp. was the most common identified organism, followed by Escherichia coli and Pseudomonas spp. Carbapenem resistance was observed in 106 (53%) strains. Almost 66% of the strains showed biofilm formation. On evaluation of associated risk factors, age (p=0.043), presence of biofilms (p=0.0001), diabetes (p=0.002), hypertension (p=0.02) and medical device use (p=0.008) had significant association, whereas sex, previous surgery and prior antibiotic use had no significant impact on the chronicity of the wound. CONCLUSION: In this study, chronicity of wounds was observed to be associated with multiple risk factors, especially the biofilm-forming ability of the strain. Biofilms are difficult to eradicate and additional measures, such as physical debridement, are important for resolving chronicity. Knowledge about specific risk factors would also allow clinicians a better understanding of the healing process and drive appropriate wound care interventions. DECLARATION OF INTEREST: A grant was received from the Indian Council of Medical Research (ICMR) for this work (grant ID: 2017-02686). The authors have no conflicts of interest to declare.

SARS-CoV-2 infection in children with chronic kidney disease.

BACKGROUND: Information on the course of SARS-CoV-2 infection in children with chronic kidney disease (CKD) is limited. METHODS: We retrospectively reviewed the presentation and outcomes of SARS-CoV-2 infection in patients with CKD followed at any of the four pediatric nephrology centers in New Delhi from April 2020 to June 2021. Outcomes, including cardiopulmonary and renal complications, were reported in relation to underlying disease category and illness severity at presentation. RESULTS: Underlying illness in 88 patients included nephrotic syndrome (50%), other CKD stages 1-4 (18.2%), CKD 5D (17%), and CKD 5T (14.8%). Thirty-two of 61 patients with symptomatic COVID-19 and 9/27 asymptomatic patients were admitted for median 10 (interquartile range 7-15) days. Seventeen (19.3%) patients developed moderate or severe COVID-19. Systemic complications, observed in 30 (34.1%), included acute kidney injury (AKI, 34.2%), COVID-19 pneumonia (15.9%), unrelated pulmonary disease (2.3%), and shock (4.5%). Nineteen (21.6%) had severe complications (AKI stage 2-3, encephalopathy, respiratory failure, shock). Eight (11%) of twelve (16.4%) patients with severe AKI required dialysis. Three (3.4%) patients, two with steroid-resistant nephrotic syndrome in relapse and one with CKD 1-4, died due to respiratory failure. Univariate logistic regression indicated that patients presenting with nephrotic syndrome in relapse or moderate to severe COVID-19 were at risk of AKI (respective odds ratio, 95%CI: 3.62, 1.01-12.99; 4.58, 1.06-19.86) and/or severe complications (respective odds ratio, 95%CI: 5.92, 1.99-17.66; 61.2, 6.99-536.01). CONCLUSIONS: Children with CKD presenting with moderate-to-severe COVID-19 or in nephrotic syndrome relapse are at risk of severe complications, including severe AKI and mortality. A higher resolution version of the Graphical abstract is available as Supplementary information.

Clinico-epidemiological profile of COVID-19 patients admitted during third wave of pandemic in a tertiary care hospital in New Delhi, India.

Even nearly two years after the first reported case, the novel coronavirus (SARS-CoV-2) continues to ebb and flow around the world. A retrospective cohort study was carried out to determine the clinico-epidemiological profile and outcome of the cases. The study analyzed secondary data from 827 patients who presented to our center with COVID-19-related illnesses between December 15, 2021, and February 15, 2022 (third wave in India). There was a significant difference in the vaccination status of patients treated at home and those admitted, with 87.9% having received two doses compared to 74% in the second group being unvaccinated. Patients who were isolated at home recovered at a rate of 99.4%, while hospitalized patients died at a rate of 26.5%. Vaccination reduces the severity of COVID-19; however, constant vigilance for new variants, precautionary measures, and increased vaccination drives are critical moving forward.   *Other members of the Safdarjung Hospital COVID-19 working group: B. Lal (Medicine), Harish Sachdeva (Anaesthesiology), Santvana Kohli (Anaesthesiology), Amandeep Jaswal (Anaesthesiology), Sumitra Bachani (Obstetrics and Gynecology), Ajay Kumar (Pediatrics), Rohit Kumar (Pulmonary Medicine), Vidya Sagar Chaturvedi (Surgery), Vinod Chaitanya (Medicine).

Evaluation of the clinical profile, laboratory parameters and outcome of two hundred COVID-19 patients from a tertiary centre in India.

COVID-19 is a pandemic with over 5 million cases worldwide. The disease has imposed a huge burden on health resources. Evaluation of clinical and epidemiological profiles of such patients can help in understanding and managing the outbreak more efficiently. This study was a prospective observational analysis of 200 diagnosed COVID-19 patients admitted to a tertiary care center from 20th march to 8th May 2020. All these patients were positive for COVID-19 by an oro-nasopharyngeal swab-rtPCR based testing. Analyses of demographic factors, clinical characteristics, comorbidities, laboratory parameters, and the outcomes were performed. The mean age of the population was 40 years with a slight male predominance (116 patients out of 200, 58%). A majority of the patients (147, 73.5 %) were symptomatic, with fever being the most common symptom (109, 54.5%), followed by cough (91, 45.5%). An older age, presence of symptoms and their duration, leukocytosis, a high quick SOFA score, a high modified SOFA score, need for ventilator support, an AST level more than 3 times the upper limit of normal (ULN), and a serum creatinine level of 2 mg/dl or greater were at a significantly higher risk of ICU admission and mortality. Presence of diabetes mellitus, AST > three times ULN, serum creatinine 2 mg/dl or higher, and a qSOFA score of 1 or higher were all associated with significantly greater odds of critical care requirement. Triage and severity assessment helps in deciding the requirement for a hospital stay and ICU admission for COVID-19 which can easily be done using clinical and laboratory parameters. A mild, moderate and severe category approach with defined criteria and treatment guidelines will help in judicious utilization of health-care resources, especially for developing countries like India.   *Other members of the Safdarjung Hospital COVID-19 working group: Balvinder Singh (Microbiology), MK Sen (Pulmonary Medicine), Shibdas Chakrabarti (Pulmonary Medicine), NK Gupta (Pulmonary medicine), AJ Mahendran (Pulmonary Medicine), Ramesh Meena (Medicine), G Usha (Anaesthesiology), Santvana Kohli (Anaesthesiology), Sahil Diwan (Anaesthesiology), Rushika Saksena (Microbiology), Vikramjeet Dutta (Microbiology), Anupam Kr Anveshi (Microbiology).

Clinical and epidemiologic profile of the initial COVID-19 patients at a tertiary care centre in India.

COVID-19 has now become a pandemic. It has spread from Wuhan, China, in December 2019 to a large number of countries within three months. The objective of this work is to report the initial experience with epidemiologic and clinical features, as well as with the management of COVID-19 patients in India. This is a descriptive case series of the first 21 COVID-19 infected patients confirmed with polymerase chain reaction (PCR) and admitted to a tertiary care centre in India from 01.02.2020 to 19.03.2020. Clinical, laboratory, and radiologic data were collected, including age, sex, nationality, travel history, symptoms, duration of stay, and comorbidities. The mean age of the population was 40.3 years with a male preponderance. Thirteen (62%) patients had recent travel history outside India in the previous 30 days, two thirds of whom had travelled to Italy. The most common symptoms were fever and cough (42.9%) followed by sore throat, headache and breathlessness. Vital and laboratory parameters were preserved in all patients and none of them required ventilatory support. Among the first 21 patients diagnosed with COVID-19 infection in India, the typical clinical presentation consisted in a mild upper respiratory tract infection predominantly affecting the young male population. One patient required supplemental oxygen. All patients recovered with no residual symptoms.   *The Safdarjung Hospital COVID 2019 working group: Nitesh Gupta, Sumita Agrawal, Pranav Ish, Suruchi Mishra, Rajni Gaind, Ganapathy Usha, Balvinder Singh, Manas Kamal Sen, Shibdas Chakrabarti (Consultant and Head, Pulmonary Medicine); NK Gupta (Professor, Pulmonary medicine); Dipak Bhattacharya (Consultant, Pulmonary medicine); Rohit Kumar (Assistant Professor, Pulmonary Medicine); Siddharth R. Yadav (Assistant Professor, Pulmonary Medicine); Rushika Saksena (Specialist, Microbiology); Rojaleen Das (Assistant Professor, Microbiology); Vikramjeet Dutta (Assistant Professor, Microbiology); Anupam Kr Anveshi (Senior Resident, Microbiology); Santvana Kohli (Assistant Professor, Anaesthesiology); Naveen KV (Assistant Professor,  Anaesthesiology); Amandeep Jaswal (Assistant Professor, Anaesthesiology).

Phenotypic and molecular characterization of Acinetobacter baumannii isolates causing lower respiratory infections among ICU patients.

BACKGROUND: Multi-drug resistant Acinetobacter baumannii has emerged as important nosocomial pathogen associated with various infections including lower respiratory tract. Limited therapeutic options contribute to increased morbidity and mortality. Acinetobacter baumannii has the ability to persist in the environment for prolonged periods. Breach in infection control practices increases the chances of cross transmission between patients and inter/intraspecies transmission of resistance elements. The present prospective work was conducted among patients with lower respiratory tract infections (LRTI) in the intensive care unit (ICU) to study the etiology with special reference to Acinetobacter baumannii and the role of immediate patient environment in the ICU as possible source of infection. Acinetobacter baumannii were characterized for antimicrobial susceptibility, mechanism of carbapenem resistance and virulence determinants. Molecular typing of the clinical and environmental isolates was undertaken to study the probable modes of transmission. MATERIALS AND METHODS: Appropriate respiratory samples from 107 patients with LRTI admitted to ICU during September 2016 to March 2017 were studied for likely bacterial pathogens. Environmental samples (n = 71) were also screened. All the samples were processed using conventional microbiological methods. Consecutive Acinetobacter spp. isolated from clinical and environmental (health care workers and environment from ICU) samples were included in the study. Antimicrobial susceptibility was performed as per CLSI guidelines. Carbapenem resistance, mediated by carbapenemase genes (blaOXA-23-like,blaOXA-24-like,blaOXA-58-like and blaNDM-1) were studied by PCR. Biofilm forming ability was tested phenotypically using microtitre plate method. Pulse Field Gel Electrophoresis (PFGE) was used to study clonality of the clinical and environmental isolates. RESULTS: The prevalence of Acinetobacter baumannii was 26.2% (28/107) and 11.26% (8/71) among patients with LRTI and environmental samples respectively. The carbapenem resistance was high, 96.42% (27/28) and 87.5% (7/8) in clinical and environmental isolates respectively. The most common carbapenemase associated with resistance was blaOXA-23-like gene followed by blaNDM-1 among both the clinical and environmental isolates. All isolates were sensitive to colistin (MIC ≤ 1 µg/ml). Biofilm production was observed among all clinical (n = 28) and 87.5% (7/8) of the environmental isolates. Line listing of the cases suggests the occurrence of infections throughout the study period with no significant clustering. On PFGE, 12 clusters were observed and 16/36 isolates were present in one single cluster that included both clinical and environmental isolates which were either carbapenem resistant or sensitive. DISCUSSION: Carbapenem resistant Acinetobacter baumannii (CRAB) is an important cause of LRTI in the ICU. PFGE suggests spread of carbapenem resistant isolates via cross transmission among patients and the environment. The detection of blaNDM-1 gene among Acinetobacter baumannii and existence of carbapenem resistant and sensitive isolates within the same clones suggests horizontal transmission of resistant genes among various bacterial species. The ability of Acinetobacter baumannii to form biofilms may contribute to its persistence in the environment. This along with breach in infection control practices are the likely factors contributing to this transmission. This information can be used to strengthen and monitor infection control (IC) and the hospital cleaning and disinfection practices to prevent spread of resistant organisms within the ICU. Colistin remains drug of choice for management of CRAB.

Linezolid resistant coagulase negative staphylococci (LRCoNS) with novel mutations causing blood stream infections (BSI) in India.

BACKGROUND: Coagulase-negative Staphylococci (CoNS) have emerged as a major causative agent of blood-stream infections (BSI). Linezolid (LZD) is currently used for treating glycopeptide and methicillin-resistant staphylococci. It is important to understand the resistance mechanism and probable transmission of LZD resistant (LR) CoNS within the hospital. METHODS: Clinically significant LRCoNS from patients with BSI were characterized using MALDI-TOF and 16S rRNA gene sequence analysis. Antimicrobial susceptibility and MIC of vancomycin and LZD were determined. LZD resistance mechanisms using PCR for the cfr gene and mutation in the V domain of the 23S rRNA gene were studied. RESULTS: The MIC of LZD ranged from 8 to 32 µg/ml. LR was observed in three different CoNS species from diverse locations within the hospital. The cfr gene was identified in all the isolates. Sequence analysis of V domain region of 23S rRNA gene confirmed mutation in single copy among 12/15 isolates with novel mutations: G2614 T and C2384T. All infections were nosocomially acquired and LZD resistance was emerging in the absence of prior LZD use. Horizontal spread of resistant isolates and cfr gene among diverse species were the probable mechanisms of transmission. CONCLUSION: The study highlights the novel mutations associated with LRCoNS and the importance of surveillance & transmission pathway within the hospital. It also systematically discusses the published information on LRCoNS.

Evaluation of an immunochromatography rapid diagnosis kit for detection of chikungunya virus antigen in India, a dengue-endemic country.

BACKGROUND: Chikungunya virus (CHIKV) and dengue virus (DENV) are arboviruses that share the same Aedes mosquito vector, and there is much overlap in endemic areas. In India, co-infection with both viruses is often reported. Clinical manifestations of Chikungunya fever is often confused with dengue fever because clinical symptoms of both infections are similar. It is, therefore, difficult to differentiate from those of other febrile illnesses, especially dengue fever. We previously developed a CHIKV antigen detection immunochromatography (IC) rapid diagnosis kit [1]. The current study examined the efficacy of previously mentioned IC kit in India, a dengue-endemic country. METHODS: Sera from 104 CHIKV-positive (by qRT-PCR) and/or IgM-positive (ELISA) subjects collected in 2016, were examined. Fifteen samples from individuals with CHIKV-negative/DENV-positive and 4 samples from healthy individuals were also examined. Of the 104 CHIKV-positive sera, 20 were co-infected with DENV. RESULTS: The sensitivity, specificity and overall agreement of the IC assay were 93.7, 95.5 and 94.3%, respectively, using qRT-PCR as a gold standard. Also, there was a strong, statistically significant positive correlation between the IC kit device score and the CHIKV RNA copy number. The IC kit detected CHIKV antigen even in DENV-co-infected patient sera and did not cross-react with DENV NS1-positive/CHIKV-negative samples. CONCLUSIONS: The results suggest that the IC kit is useful for rapid diagnosis of CHIKV in endemic areas in which both CHIKV and DENV are circulating.

Clinical, Serological, and Virological Analysis of 572 Chikungunya Patients From 2010 to 2013 in India.

Background: Chikungunya fever (CHIK) is a major public health concern in India. Characterized by acute fever with joint pain and swelling, most patients recover from this self-limiting illness in 7-10 days, with cessation of joint pain post-acute episode. However, in some patients, joint pain persists, lasting for months or even years. The precise correlates to the chronic phase of this debilitating illness and/or this remarkable heterogeneity in disease manifestation are poorly understood. Methods: We evaluated 572 chikungunya patients from India who were recruited on the basis of positive real-time polymerase chain reaction and/or CHIK virus immunoglobulin (IgM) after receiving consent. Arthralgic conditions were monitored using visual analog score (VAS) 12 weeks after onset of fever in 130 patients. Initial viral load, IgG, and initial neutralization response were assayed and correlated with clinical and VAS information in 40 patients. Results: Our extensive screening revealed that patients with higher initial viral loads during the acute phase of illness had poor prognosis at the post-acute phase with more restricted joint movement and higher VAS. Additionally, patients who showed early seroconversion to neutralizing IgG responses had better prognosis, as many of these patients did not manifest restricted joint movements at the post-acute phase. Conclusions: Our study sheds light on chikungunya disease with respect to disease progression and assesses clinical, virological, and serological parameters of chikungunya disease severity. Importantly, it reveals that initial high viral load and neutralizing IgG response may function in a seemingly contrasting manner to negatively or positively dictate disease outcome.

Risk factors for fecal carriage of carbapenemase producing Enterobacteriaceae among intensive care unit patients from a tertiary care center in India.

BACKGROUND: Resistance amongst the commensal flora is a serious threat because a very highly populated ecosystem like the gut, may at a later stage, be a source of extra intestinal infections, resistant strains may spread to other host or transfer genetic resistance element to other members of micro-biota including pathogens. This study was carried out to assess fecal colonization by carbapenemase producing Enterobacteriaceae (CPE) and associated risk factors among 100 patients admitted to intensive care unit (ICU). The phenotypic and molecular characterizations of CPE were also included. RESULTS: Colonization with CPE was observed in 6.6 % (8/122) controls. Among ICU patients, fecal carriage of CPE was significantly higher on day 4 (D4) (22 %) as compared to day 1 (D1) (11 %) (p value 0.002). The carbapenemase genes detected included OXA- 48, 181, KPC and NDM-1 with NDM-1 being the predominant carbapenemase in both ICU D1 and D4. Among the 50 CPE isolates, 8 (16 %) were susceptible to meropenem and imipenem (Minimum inhibitory concentration; MIC ≤ 1 mg/L) and all were susceptible to colistin (MIC range 0.125 - 1 mg/L) and tigecycline (MIC range 0.06- 1.5 mg/L). The risk factors associated with CPE carriage were duration of ICU stay, use of ventilator and aminoglycosides. CONCLUSIONS: Prior colonization with CPE could result in their influx and spread in ICU, challenging infection control measures. Exposure to ICU further increases risk of colonization with diverse carbapenemase-producing Enterobacteriaceae. Gut colonization with these strains may be a source of endogenous infection and horizontal transfer of these genes in future.

Hemozoin Pigment: An Important Tool for Low Parasitemic Malarial Diagnosis.

Low parasitemic condition in malaria remains a diagnostic challenge; as the available diagnostic methods failed to detect. Currently, hemozoin (Hz) pigment is gaining attention in the diagnosis of malaria. The major drawback is ease of detection of Hz in routine practice. A pilot study was conducted to evaluate the role of Hz pigment and to compare the performance of quantitative buffy coat assay (QBC) and PCR in such conditions. Clinically suspected cases of malaria were examined by both Giemsa stain and immunochromatographic test (ICT). Samples positive by ICT and negative by Giemsa stain were further examined by nested PCR targeting 18S rRNA and QBC for the presence of malaria parasites and pigments. Thirty blood samples fulfilled the inclusion criteria out of which 23 were Plasmodium vivax (Pv), 4 Plasmodium falciparum (Pf), and 3 mixed (Pv and Pf) by immunochromatographic test. Twenty-one out of 30 (70%) were positive by nested PCR in comparison to 25/30 (83%) by QBC. Samples containing both malaria parasites and Hz pigment by QBC completely showed concordance with the PCR result. However, 61% of total samples containing only Hz pigment were observed positive by PCR. Hz pigment remains an important tool for malaria diagnosis. Identification of leukocytes containing pigments by QBC not only indicates recent malarial infections but also puts light on severity of the disease. QBC assay is a rapid, highly sensitive, and cost-effective method to detect malaria parasites and Hz pigment especially in low parasitemic conditions.

Clinically and microbiologically derived azithromycin susceptibility breakpoints for Salmonella enterica serovars Typhi and Paratyphi A.

Azithromycin is an effective treatment for uncomplicated infections with Salmonella enterica serovar Typhi and serovar Paratyphi A (enteric fever), but there are no clinically validated MIC and disk zone size interpretative guidelines. We studied individual patient data from three randomized controlled trials (RCTs) of antimicrobial treatment in enteric fever in Vietnam, with azithromycin used in one treatment arm, to determine the relationship between azithromycin treatment response and the azithromycin MIC of the infecting isolate. We additionally compared the azithromycin MIC and the disk susceptibility zone sizes of 1,640 S. Typhi and S. Paratyphi A clinical isolates collected from seven Asian countries. In the RCTs, 214 patients who were treated with azithromycin at a dose of 10 to 20 mg/ml for 5 to 7 days were analyzed. Treatment was successful in 195 of 214 (91%) patients, with no significant difference in response (cure rate, fever clearance time) with MICs ranging from 4 to 16 µg/ml. The proportion of Asian enteric fever isolates with an MIC of ≤ 16 µg/ml was 1,452/1,460 (99.5%; 95% confidence interval [CI], 98.9 to 99.7) for S. Typhi and 207/240 (86.3%; 95% CI, 81.2 to 90.3) (P < 0.001) for S. Paratyphi A. A zone size of ≥ 13 mm to a 5-µg azithromycin disk identified S. Typhi isolates with an MIC of ≤ 16 µg/ml with a sensitivity of 99.7%. An azithromycin MIC of ≤ 16 µg/ml or disk inhibition zone size of ≥ 13 mm enabled the detection of susceptible S. Typhi isolates that respond to azithromycin treatment. Further work is needed to define the response to treatment in S. Typhi isolates with an azithromycin MIC of >16 µg/ml and to determine MIC and disk breakpoints for S. Paratyphi A.

Polydispersity-mediated high efficacy of an in-situ aqueous nanosuspension of PPEF.3HCl in methicillin resistant Staphylococcus aureus sepsis model.

Recently, World Health Organization declared antimicrobial resistance as the third greatest threat to human health. Absence of known cross-resistance, new class, new target, and a new mode of action are few major strategies being undertaken by researches to combat multidrug resistant pathogen. PPEF.3HCl, a bisbenzimidazole was developed as highly potent antibacterial agent against ESKAPE (Enterococcus faecium, Staphylococcus aureus, Klebsiella pneumoniae, Acinetobacter baumannii, Pseudomonas aeruginosa, and Enterobacter species) pathogens, targeting topoisomerase IA. The present work encompasses a radical on-site generation of In-situ nanosuspension of PPEF.3HCl with enhanced efficacy against methicillin resistant S. aureus in septicemia model. We have generated instantaneously a PPEF.3HCl nanosuspension (IsPPEF.3HCl-NS) by mixing optimized monophasic PPEF.3HCl preconcentrate in propylene glycol into an aqueous medium comprising tween 80 as stabilizer. The IsPPEF.3HCl-NS showed precipitation efficiency of > 90 %, average particle size < 500 nm, retained upto 5 h, a negative zeta potential and bi/trimodal particle size distribution. Differential scanning calorimetry, X-ray diffraction confirmed partial amorphization and transmission electron microscopy revealed spherical particles. IsPPEF.3HCl-NS was non-hemolytic and exhibited good stability in serum. More significantly, it exhibited a âˆ¼ 1.6-fold increase in macrophage uptake compared to free PPEF.3HCl in the RAW 264.7 macrophage cell line. Confocal microscopy revealed accumulation of IsPPEF.3HCl-NS within the lysosomal compartment and cell cytosol, proposing high efficacy. In terms of antimicrobial efficacy, IsPPEF.3HCl-NS outperforms free PPEF.3HCl against clinical methicillin sensitive and resistant S. aureus strains. In a pivotal experiment, IsPPEF.3HCl-NS exhibited over 83 % survival at 8 mg/kg.bw and an impressive reduction of âˆ¼ 4-5 log-fold in bacterial load, primarily in the kidney, liver and spleen of septicemia mice. IsPPEF.3HCl-NS prepared by the In-situ approach, coupled with enhanced intramacrophage delivery and superior efficacy, positions IsPPEF.3HCl-NS as a pioneering and highly promising formulation in the battle against antimicrobial resistance.

Application of LAMP assay for detection of carbapenem-resistant Acinetobacter calcoaceticus-Acinetobacter baumannii complex in ICU admitted sepsis patients: A rapid and cost-effective diagnostic tool.

Carbapenem-resistant significant members of Acinetobacter calcoaceticus-Acinetobacter baumannii (CR-SM-ACB) complex have emerged as an important cause of sepsis, especially in ICUs. This study demonstrates the application of loop-mediated-isothermal-amplification (LAMP) assay for detection of CR-SM-ACB-complex from patients with sepsis. Whole-blood and culture-broths(CB) collected from patients with culture-positive sepsis were subjected to LAMP and compared with PCR, and RealAmp. Vitek-2 system and conventional PCR results were used as confirmatory references. The sensitivity and specificity of LAMP(97 % & 100 %) and RealAmp(100 % & 100 %) for detection of CR-SM-ACB-complex from CB were better than PCR(87 % & 100 %). Diagnostic accuracy of LAMP, RealAmp, and PCR for detection of SM-ACB-complex from CB was 98.5 %, 100 %, and 88.5 % respectively. Turnaround time of Culture, LAMP, PCR, and RealAmp was 28-53, 6-20, 9-23, and 6-20hours, respectively. LAMP is a simple, inexpensive tool that can be applied directly to positive CB and may be customized to detect emerging pathogens and locally-prevalent resistance genes and to optimize antimicrobial use.

Community acquisition of ß-lactamase producing Enterobacteriaceae in neonatal gut.

BACKGROUND: Commensal flora constitutes a reservoir of antibiotic resistance. The increasing variety of ß-lactamases and the emergence of Carbapenem resistant Enterobacteriaceae (CRE) in community, raise concerns regarding efficacy of ß-lactams. It is important to know the exact load of antibiotic resistance in the absence of any antibiotic selection pressure including via food and water.In the present study gut colonization in neonates with no direct antibiotic pressure was used as a model to evaluate ß-lactam resistance in the community. RESULTS: In this prospective study, 75 healthy, vaginally delivered, antibiotic naive, breast fed neonates were studied for gut colonization by Extended spectrum ß-lactamases (ESBL), AmpC ß-lactamases hyperproducing Enterobacteriaceae and CRE on day 0, 21 and 60. Total 267 Enterobacteriaceae were isolated and E.coli was the predominant flora. ESBL, AmpC and coproduction was seen in 20.6%, 19.9% and 11.2% isolates respectively. ESBL carriage increased threefold from day 1 to 60 showing predominance of CTX-M group 15 (82.5%), ampC genes were heterogeneous. Colonization with CRE was rare, only one baby harboured Enterobacter sp positive for kpc-2. The reservoirs for these genes are likely to be mother and the environment. CONCLUSIONS: Data strongly suggests that in absence of any antibiotic pressure there is tremendous load of antibiotic resistance to ß-lactam drugs. Wide spread presence of ESBL and AmpC can drive rapid emergence and dissemination of CRE. This is the first report from India which depicts the smaller picture of true antibiotic pressure present in the Indian community.

Neonatal septicaemia caused by diverse clones of Klebsiella pneumoniae & Escherichia coli harbouring blaCTX-M-15.

BACKGROUND & OBJECTIVES: Information about the genetic diversity of the extended-spectrum ß-lactamases (ESBLs) and the clonal relationship of the organisms causing neonatal infections is limited, particularly from India where neonatal mortality is high. This study was undertaken to investigate the molecular epidemiology and risk factors associated with neonatal septicaemia caused by ESBL-producing Klebsiella pneumoniae and Escherichia coli. METHODS: Bloodstream isolates (n=26) of K. pneumoniae (n=10) and E. coli (n=16) from the neonates admitted in a tertiary care hospital in New Delhi during January to May 2008 were characterized. Antimicrobial susceptibility tests were carried out and ESBL production was assessed phenotypically. PCR was carried out for ESBL and ampC genes. Genotyping was performed by pulsed-field gel electrophoresis (PFGE). Conjugation experiments were done to determine the mobility of ESBL genes. Risk factors associated with ESBL-producing K. pneumoniae and E. coli infections were analysed. RESULTS: Resistance rates to most of the antibiotics tested were high, except for imipenem. Among the isolates tested, 60 per cent of K. pneumoniae and 75 per cent of E. coli were ESBL producers. PFGE of the isolates demonstrated a vast diversity of genotypes with no epidemic clones. Despite the clonal diversity, blaCTX-M-15 was detected in 100 per cent of ESBL-positive isolates. The other genes present in ESBL-positive isolates were blaTEM-1, blaSHV-1 , blaSHV-28 , blaSHV-11 , and blaSHV-12 . Class 1 integrons were detected in 7 of 18 ESBL-positive isolates. Moreover, the plasmid carrying blaCTX-M-15 , in E. coli and K. pneumoniae were self transferable. Feeding through an enteral tube was identified as the only risk factor for sepsis by ESBL-producing organisms. INTERPRETATION & CONCLUSIONS: The study emphasises the presence of blaCTX-M-15 in clonally diverse isolates indicating probable horizontal transfer of this gene. The widespread dissemination of CTX-M-15 is of great concern as it further confines the limited therapeutic interventions available for neonates.

Bacteraemic Haemophilus influenzae type B pneumonia complicated with empyema--report of a case and review of literature.

A case of bacteraemic pneumonia complicated with pleural empyema due to Haemophilus influenzae type b is reported in a one-year old previously healthy child who had apparently no other associated medical condition. The organism was isolated from both the pleural fluid aspirate and blood of the patient with pneumonia. She was successfully treated with parenteral ampicillin and chloramphenicol alongwith intercostal chest tube drainage. The case is notable because it adds to the existing disease spectrum of invasive Hib diseases and brings awareness to the existing burden of the disease in Asia. In addition, it reflects the urgent need to include Hib vaccine in the current immunization program in India.