Blood culture remains the most important microbiological investigation in the management of sepsis. The conventional definition of sepsis has been challenged recently and likely to increase number of patients screened for sepsis due to the change in the definition. Blood culture contamination still remains a challenge, especially in resource poor settings where educational facilities are limited. Clinical correlation of positive (as well as negative) blood culture is an important aspect of the investigation where a clinical microbiologist can significantly impact on the management of sepsis.

Introduction: Emerging multidrug resistance continues to be a major concern in healthcare settings. The aim of the study was to determine the resistance pattern of multidrug-resistant (MDR) Enterobacteriaceae causing urinary tract infections in our hospital and to report the occurrence of extended-spectrum beta-lactamase (ESBL), AmpC and metallo-beta-lactamase (MBL) production in them. Materials and Methods: Out of 280 MDR strains collected over a period of one year, 130 strains of Escherichia coli (96), Klebsiella spp. (31) and Enterobacter spp. (3) resistant to the second- and third-generation Cephalosporins were selected for further testing. Cefotaxime, Cefotaxime-Clavulanic acid, Ceftazidime, Ceftazidime-Clavulanic acid and Cefepime, Cefepime-Clavulanic acid Etest strips, Cefoxitin and Cefotetan with Boronic acid and Imipenem/Imipenem-EDTA Etest strips were used to detect ESBLs, AmpC and MBLs. Multiplex polymerase chain reaction (PCR) was done to detect plasmid-mediated AmpC genes. Results: Among 130 Cefoxitin-resistant strains, Cefoxitin-Boronic acid inhibitor method detected AmpC phenotype in 116 (89.2%) isolates. The overall occurrence of AmpC (n = 280) was 116 (41.42%). 92 (32.8%) isolates were found to be ESBL producers by the Clinical and Laboratory Standards Institute confirmatory method. ESBL production was detected in 107 (38.2%) more isolates by Cefepime/Cefepime-Clavulanic acid Etest. MBL producers were relatively low in our study 5 (1.8%). PCR detected CIT genotype (CMY-2) in 13 isolates (4.6%). Conclusion: This study reveals high prevalence of AmpC and ESBL co-carriage suggesting plasmid-mediated spread, indicates the need for surveillance of resistance mechanisms and takes necessary measures to control the emergence of MDR organisms.

Background and Objectives: Surgical site infection (SSI) continues to be a major healthcare-associated infection. The objective of the study was to find the bacterial profile of SSI occurring within one week of surgery and to determine their antibiotic sensitivity pattern in a tertiary care centre. Materials and Methods: Pus from the surgical sites was cultured, the organisms were identified and their antibiotic sensitivity was found by conventional methods. Data were collected to assess the risk factors. The infection rate was compared with the available standards. Results: During the one-year study period, 2472 post-operative patients were followed up, of whom 227 got infected. The infection rate was 9.2% which was high compared to the Centers for Disease Control statistics of 1.9%. The predominant isolate was Staphylococcus aureus 50 (28.2%), followed by Escherichia coli 48 (27.1%). Interpretation and Conclusion: Inpatients were studied for the first one week only because this was the optimum period for contracting infections. Moreover, majority of the patients got discharged from the hospital after one week. S. aureus was the predominant pathogen in Class I wounds, and E. coli was predominant in Class III wounds.

Context: Bloodstream infections (BSIs) caused by yeasts have an increasing frequency due to the growing population of immunosuppressed individuals. Among yeasts, Candida remains the most prevalent species with the increase in the incidence of non-albicans Candida species. Apart from Candida, other yeasts are also involved in causing BSI. High mortality associated with Candida and other yeast infection can be reduced by prompt and appropriate antifungal therapy. Hence, rapid identification and speciation of yeasts isolated from blood play a significant role in the management of the patients. Since conventional methods used for speciation of Candida and other yeasts are laborious, time-consuming and often unclear, rapid and accurate molecular techniques are required. Materials and Methods: Instead of using purified genomic DNA as template for polymerase chain reaction (PCR), we used yeast colony and cell suspensions in water and 0.10M potassium hydroxide as template for PCR. Candida albicans, Trichosporon and Cryptococcus neoformans were used as reference strains. Further, a total of 100 yeast isolates were also tested. All reactions were performed using the universal fungal primers ITS1 and ITS4; the PCR products were then digested with restriction enzyme (Msp1). Results: Direct colony PCR (DCPCR) produced sharp and distinct bands compared to the cell suspensions with the reference strains. All the 100 clinical isolates tested also produced distinct bands. Conclusion: DCPCR approach not only reduces the DNA template preparation time but is also easy, rapid and reduces the cost of PCR.

Background: In June 2013, flash floods caused a great loss of human life and infrastructure in the Sub-Himalayan region of Uttarakhand, India. An outbreak of scrub typhus caused by Orientia tsutsugamushi occurred in the district of Chamoli, Rudraprayag and Pauri Garhwal after the disaster. The present study was conducted with the objective to describe this outbreak in terms of time, place, person and clinical features and to compare the outbreak with the pre-disaster status of the area. Materials and Methods: This study was conducted in Veer Chandra Singh Garhwali Government Medical Science and Research Institute, Srinagar, Pauri Garhwal from June to December 2013. Study subjects were patients from disaster-affected areas. Definition criteria were used for clinically suspected, probable and confirmed cases of scrub typhus. All the samples were subjected to immunochromatographic test out of which 229 were confirmed by ELISA. Results: A total of 283 samples of patients with undiagnosed fever were tested and 229 (80.9%) showed the presence of IgM antibody by ELISA against scrub typhus. The maximum number of cases (213) were found between July and November and were mainly confined to the districts mentioned above. The main clinical features were gastrointestinal symptoms (53%), rash (51%), myalgia (71%), acute respiratory distress syndrome (ARDS) (2%), hepatorenal syndrome (1.7%), coagulopathy (18%) and eschar occurred only in five cases (2%). Conclusions: An increase in the number of cases of scrub typhus was observed after the floods in Uttarakhand, which suggests that a post-disaster epidemic had occurred. Scrub typhus should also be considered in the differential diagnosis of acute febrile illness with gastrointestinal symptoms, rash, myalgia, ARDS, hepatorenal syndrome and coagulopathy. Eschar being pathognomonic, may not always be seen, and its absence does not rule out scrub typhus.

Background: Infections caused by carbapenem-resistant Gram-negative bacteria are a cause for concern due to the limited choice of antibiotics available for their treatment. Aims, Settings and Design: This study screened multidrug-resistant (MDR) Gram-negative bacilli isolated from clinical samples over a period of one year, for carbapenem resistance and characterised them using phenotypic methods such as combined disc diffusion test (CDDT), modified Hodge test (MHT), E-test for metallo-beta-lactamase (MBL) and molecular method, PCR. Materials and Methods: Two hundred and ten MDR Gram-negative bacilli were screened for carbapenem resistance using Imipenem and Meropenem disc diffusion. These were further checked for carbapenemase production by CDDT, MHT and E-test for MBL. Those positive by E-test were subjected to PCR. Uniplex PCR for New Delhi metallo-beta-lactamase-1 was used for Escherichia coli and Klebsiella pneumoniae, and multiplex PCR for Imipenemase and Verona imipenemase was used for Pseudomonas aeruginosa and Acinetobacter baumannii isolates. Results: Twenty-three (11%) isolates were found to be carbapenem-resistant and included E. coli (six) K. pneumoniae (three), P. aeruginosa (five) and A. baumannii (nine). Seventeen (74%) isolates were positive by phenotypic methods and were subjected to PCR. Out of eight Enterobacteriaceae isolates subjected to PCR, all were positive for bla_NDM gene. All were negative for bla_KPC gene. All five A. baumannii isolates subjected to PCR were found to contain bla_VIM gene. Two out of four P. aeruginosa isolates were positive for bla_IMP, one was positive for bla_VIM gene. One P. aeruginosa isolate was positive for both bla_IMP and bla_VIM gene. Conclusions: In view of the increasing resistance of Gram-negative bacilli to carbapenems, rational use of antibiotics needs to be emphasised.

Background: Helicobacter pylori (HP) is the most common bacterial infection in humans. It affects > 50% of world population and remains the major aetiological agent of ailments of upper gastrointestinal (GI) tract of humans. HP eradication prevents pre-neoplastic changes of gastric mucosa and regression of gastric mucosa-associated lymphoid type lymphoma. For these reasons and many others, it is worthwhile testing patients with dyspepsia for HP, who might benefit from eradication therapy. Objective: To study the prevalence of HP infection among dyspeptic patients in a tertiary care centre in South Kerala. Materials and Methods: Gastric biopsy specimens were collected from referred patients with dyspepsia from Southern districts of Kerala (Thiruvananthapuram, Kollam and Pathanamthitta) attending the Gastroenterology Outpatient Department, who underwent upper GI endoscopy, during a one-year period from December 2009 to November 2010. The serum samples were collected for doing HP IgG enzyme-linked immunosorbent assay test. The biopsy specimens were used for doing rapid urease test (RUT) and culture was done in patients showing urease positivity and/or high level of IgG antibody titre. Results: The study was conducted on 250 consecutive dyspeptic patients, with an age range of 7-73 years. Significant IgG titres (>37.5 IU/ml) were detected in 89 cases (35.6%), while the RUT was positive in 30 cases (12%). Subjects were considered positive for HP infection by combined IgG antibody and/or urease test positivity and negative when both the tests were negative. In this study, ninety patients (36%) were positive for HP infection, and 160 (64%) were negative. Culture was done in patients showing urease positivity and/or high level of IgG antibody titre and proportion of culture positivity among this group was 34%. Conclusion: Seroprevalence of HP infection in South Kerala was found to be low (35.6%). Low infection rates were seen in children, similar to that in developed countries, and it is likely that the prevalence of HP may fall in the coming years. Due to the lower percentage of HP infection in this area, a test and treat policy is applicable as in many developed countries.

Background: Fungal infections constitute a major health problem all over the world. Signs and symptoms induced by various dermatophytic infections are hardly distinguishable clinically from each other. Hence, characterisation by in vitro culture is required for the appropriate diagnosis and treatment as well as for studying the epidemiological characteristics in a region. Objectives: The objectives of this study are: (1) To isolate and identify dermatophytes affecting skin and nail. (2) To compare two different culture media, namely Sabouraud's dextrose agar (SDA, Himedia Laboratories, Mumbai) with Chloramphenicol and Actidione with dermatophyte test medium (DTM, Hi-Media Laboratories). Materials and Methods: This is a cross-sectional study in which patients attending the outpatient wing of the Department of Dermatology and Venereology, Government Medical College, Thiruvananthapuram, Kerala, India, with clinical features of dermatophytosis were included from March 2011 to February 2012. Skin and nail scrapings were subjected to direct microscopy by 10% potassium hydroxide (KOH), 40% KOH and cultured on SDA with Actidione (Hi-Media Laboratories) and DTM (Hi-Media Laboratories). Results: The total number of samples in this period was 150, of which 99 (66%) samples were positive by direct microscopy and 74 (49.33%) were positive by culture. The most common clinical type was tinea corporis 75 (50%) followed by tinea cruris 40 (26.67%). Out of the 74 isolates, Trichophyton rubrum 40 (54.05%) was the most common species followed by Trichophyton mentagrophytes 29 (39.19%), Microsporum gypseum three (4.05%), Trichophyton schoenleinii one (1.35%) and Epidermophyton floccosum one (1.35%).Nearly 86.1% of the dermatophytes were isolated on DTM within 5-10 days of inoculation whereas 47.05% were isolated on SDA within 10 days of inoculation. This was statistically significant with P < 0.0001 (χ² = 22.43). Conclusion: DTM can be used as a rapid screening medium for the isolation and identification of dermatophytes compared to SDA with Actidione. However, DTM is inferior to SDA with Actidione in the identification of dermatophyte species.

Cryptococcus neoformans and Herpes simplex virus (HSV) are known agents of fungal and viral meningitis, respectively, in immunocompromised patients but rarely seen in immunocompetent patients. The present report is of an immunocompetent patient, engineer by profession, who was diagnosed to have meningitis with two organisms. The patient presented with fever, chills and headache for 25 days, altered sensorium for four days and altered speech for two days. Lumbar puncture was done, cerebrospinal fluid was sent for serology and culture was positive for budding yeast cells with capsule in India ink preparation. Culture yielded mucoid, cream-coloured colonies, and latex agglutination for Cryptococcus antigen was positive. Polymerase chain reaction on CSF was positive for HSV-1. The patient was treated with Ceftriaxone, Doxycycline, Acyclovir, injection Mannitol and Amphotericin B.

Even though Staphylococcus aureus is present as normal flora on the skin, it is a pathogen when present in otherwise normally sterile body fluids such as cerebrospinal fluid and blood. Methicillin-resistant S. aureus (MRSA), presumed to originate from a patient or carrier, is responsible for both community-acquired as well as hospital-acquired infections (HAIs). Coagulase-negative staphylococci (CoNS) are present as normal flora on skin but cause vegetations on valve leaflets in subacute bacterial endocarditis and surgical site infection. MRCoNS arises from the skin flora of hospitalised patients and can contribute to similar infections in the hospital setting. We present three cases of Methicillin-resistant staphylococci causing HAI, MRSA causing meningitis with obstructive hydrocephalus, MRCoNS causing bacteraemia in a child with congenital cyanotic heart disease and MRCoNS causing surgical site infection on an amputation stump.

Neisseria meningitidis is one of the primary pathogens of pyogenic meningitis and has the potential to cause large epidemics. There are 13 serogroups of N. meningitidis that have been identified, six of which (A, B, C, W, X and Y) can cause epidemics. We report a case of acute pyogenic meningitis caused by N. meningitidis in a six-month-old male child who presented with fever of four-day duration and irritable cry for one day. N. meningitidis was isolated from his blood using automated blood culture system. Latex agglutination test for N. meningitidis antigen was positive from cerebrospinal fluid. The baby responded well to Ceftriaxone but developed subdural effusion.

Histoplasmosis is a very rare disease in non-endemic areas. It is a granulomatous disease caused by a saprophytic dimorphic fungus Histoplasma capsulatum. In immunocompromised individuals, it may cause a progressive and potentially fatal disseminated disease. Isolated cutaneous histoplasmosis is rare in renal transplant recipients. We report a case of a 45-year-old male who presented with skin lesions 4 years after renal transplantation. Histopathological examination of biopsy taken from the skin lesions showed intracellular budding yeast cells and was confirmed by culture of the biopsy sample. The patient was treated with Itraconazole and responded well with disappearance of all skin lesions.

Background: Group B Streptococcus (GBS) has reemerged as a major pathogen during the past few decades. Newborns with early-onset GBS disease acquire infection from the maternal genital tract. The aim of the present study was to find the prevalence of GBS among antenatal cases and to evaluate the conventional and CHROMagar^TM Strep B agar method in the detection of GBS colonization among pregnant women. Materials and Methods: A total of 160 vaginal swabs were collected from pregnant women of 35-37 weeks of gestation and inoculated onto 5% sheep blood agar and CHROMagar^TM Strep B agar. GBS grown on 5% sheep blood agar and CHROMagar^TM Strep B agar were confirmed by biochemical and latex agglutination tests. Results: GBS was detected in 14.38% of pregnant women. CHROMagar^TM Strep B agarshowed 100% sensitivity and specificity in comparison with the conventional method. Conclusion: In the present study, GBS was prevalent in 14.38% of the antenatal cases. CHROMagar^TM Strep B agar with 100% sensitivity and specificity can be used to screen all pregnant women for GBS colonization as it does not require expertise in identification.

Purpose: For antibiotic susceptibility results, conventional culture and sensitivity methods take 48 h after a blood culture are flagged positive by automated systems. Early initiation of targeted antibiotic therapy is essential for effective management of sepsis to reduce morbidity, mortality, cost of treatment and prevent antibiotic resistance. The objective of this study was to evaluate direct sensitivity test (DST) as a potential tool to get reliable antibiotic susceptibility results 24 h earlier. Materials and Methods: Blood cultures that flagged positive from 1^st January 2015 to 30^th June 2015 by BacT/ALERT were stained by Gram stain. All blood cultures with only one kind of Gram-negative bacteria by Gram stain were simultaneously cultured and sensitivity tests put up directly (DST) from the broth using disk-diffusion method according to the British Society of Antimicrobial Chemotherapy guidelines. DST results available next day were compared with conventional antibiotic susceptibility test (AST). Results of DST (test method) and AST (reference method) were compared for agreements or errors. Results: Of the fifty Gram-negative isolates tested, we observed 91.5% categorical agreement or no error (κ = 0.407, P < 0.001). Conclusions: DST using disk diffusion from positive blood culture broths helps initiate early targeted antibiotic therapy. There is a high concordance between DST and AST.

Context: Water is an important resource for hospitals. There are few studies about the quality, quantity and cost of water that is required for health care. Aims: To study the quality, quantity, cost and applications of water in a hospital. Settings and Design: Observational study. The study was done in a cancer hospital in eastern India. Methods and Material: Water generation and consumption patterns and costing were assessed after: Discussion with the engineers; analysis of documented records; observation of patient/visitor/staff behaviours; measurement of flow rates and metered readings; Individual water consumption surveys. Statistical Analysis Used: None. Results: The total filtered reverse osmosis (RO) water used by the hospital per day was 200,000 L. This equated to 1093 L/patient/day. The volume of filtered reversed osmosis water consumed showed that the total water usage for drinking was 1%, water usage for hand-washing was 18%, water usage for showering was 6%, kitchen water consumption was 2%, housekeeping activities usage was 4%, central sterile supply department usage was 4%, heating, ventilation, and air conditioning systems usage was 36%, hot water consumption was 5% and toilet flush usage was 24%. Cost was Rs. 1119/- for 10,000 liters of RO water and about Rs. 31/- for 10,000 liters of raw water. Conclusions: The economics of hospital water both in terms of consumption and cost is a valuable source of information for hospital planners, administrators and hospital engineers. Key Messages: • Water is a precious resource. Wastage needs to be minimized • It is important for hospitals to monitor its water usage • Safe and adequate water is essential for effective hospital infection control and monitoring its microbiological quality is of paramount importance