Inconclusive SARS CoV-2 reverse transcription-polymerase chain reaction (RT-PCR) reports for the detection of infection in symptomatic patients or during screening of asymptomatic contacts can cause clinical, diagnostic and infection control uncertainty. It has been noted that up to 5% of COVID RT-PCR reports may be inconclusive. The reasons for these inconclusive reports are varied and may be classified into virological causes, sample collection and sample quality-related issues and finally technical issues related primarily due to problems in RNA extraction. The objective of this article is to discuss these causes and suggest corrective measures.

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Chronic suppurative otitis media is an infection of the middle ear and mastoid cavity. It is one of the most common causes of deafness. Low social-economic status, overcrowding, malnutrition and poor hygiene are the common risk factors and it commonly affects infants and children. The common pathogens responsible are Pseudomonas aeruginosa, Staphylococcus aureus, Escherichia coli, Aspergillus species, Candida species etc. Several studies similar to this have been done previously from various regions of our country and also from several parts of the world. There are no studies done recently from this region to determine the bacteriological profile of aerobic bacteria and their antibiotic susceptibility in this region. The study was done for six months and the sample size was 80. Ear swab culture was done on blood agar, MacConkey agar, and chocolate agar. The culture plates were processed and identification of the isolates and antibiotic susceptibility were done as per standard methods. Samples with bacterial growth were 66 (82.5%), fungal growth were 10 (12.5%) and 4 (5%) samples had no growth. There were 82 isolates of bacteria and fungi from 80 patients. The predominant bacteria were Pseudomonas aeruginosa with 31 (37.80%) isolates followed by Staphylococcus aureus 25 (30.49%). All 31(100%) P. aeruginosa isolates were sensitive to colistin and polymixin B, while 30 (96.8%) isolates showed sensitivity to piperacillin-tazobactam, cefoperazone-sulbactam and aztreonam. Only 21 (67.7%) isolates were sensitive to gentamicin, 23 (74.2%) isolates were sensitive to ciprofloxacin, whereas 24 (77.4%) isolates were sensitive to levofloxacin. All 25 (100%) S. aureus isolates showed sensitivity to vancomycin, teicoplanin and linezolid whereas 23 (92%) isolates showed sensitivity to amikacin, doxycycline and clindamycin. Only one isolate (4%) was sensitive to ampicillin whereas 8 (32%) were sensitive to amoxicillin-clavulanic acid in vitro. P. aeruginosa and S. aureus are the most common aerobic bacterial pathogens in chronic suppurative otitis media. P. aeruginosa showed 100% sensitivity to Colistin and Polymixin B followed by Piperacillin-tazobactam (96.8%), Cefoperazone-sulbactam (96.8%) and Aztreonam (96.8%). S. aureus showed 100% sensitivity to Vancomycin, Teicoplanin and Linezolid followed by Amikacin, Doxycycline and Clindamycin (92% each). Culture and sensitivity should be done in chronic suppurative otitis media before initiating antibiotic therapy.

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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.

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Mucormycosis is a life-threatening invasive fungal infection usually occurring in immunocompromised patients or in patients with uncontrolled diabetes mellitus. It carries a high mortality rate. Aspergillosis which is caused by Aspergillus species may cause an invasive disease in immunosuppressed patients. A 63-year-old female patient with type 2 diabetes mellitus presented with sudden onset painless diminution of vision in the right eye and restriction of orbital movements. Oculo-rhino-cerebral mucormycosis was diagnosed on contrast-enhanced computed tomography scan. Histopathology and fungal culture showed mixed infection with Mucor, Aspergillus and Candida. Liposomal Amphotericin B was started and surgical debridement was done.

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Talaromyces marneffei, earlier called Penicillium marneffei, is a fungus which causes infection mostly in immunocompromised patients. Here, we report a case of T. marneffei causing chronic skin infection in a farmer who has no history of immune deficiency. Antifungal treatment was started, but the patient expired due to complications of deep vein thrombosis.

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CONTEXT: Rapid detection of pathogens in infected body fluids will help in establishing an aetiological diagnosis, thereby facilitating specific therapy. Molecular methods have an advantage over conventional bacteriological techniques in particularly identifying slow-growing, fastidious or non-cultivable organisms. AIM: The aim of this study is to evaluate the use of 16S rRNA polymerase chain reaction (PCR) for detecting bacterial pathogens in body fluids and compare with conventional culture methods. SETTINGS AND DESIGN: This study was done at the Clinical Laboratory of the Department of Microbiology. This was a cross-sectional study design. MATERIALS AND METHODS: A total of 100 consecutive samples which included synovial fluid, cerebrospinal fluid, ascitic fluid and pleural fluid received in the laboratory during the study period were subjected to PCR for 16S rRNA using specific primers and conventional culture by standard protocol. Samples which were positive for 16S rRNA were sequenced to identify the organism. Results of sequenced products were compared in terms of number of organisms, with culture isolates. RESULTS: The detection rate of 16S rRNA PCR was at 13% as compared to culture at 3% (P = 0.0009). The diagnostic sensitivity and specificity of the PCR were 100% and 89.7%, respectively. The concordance of PCR and culture for both identical positive and negative samples was 90%. CONCLUSIONS: The 16S rRNA PCR proved to be rapid method for detection of bacterial pathogens in body fluids. It may be a valuable tool in the diagnostic armamentarium for differentiating bacterial infection from others and starting empiric treatment.

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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.

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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.

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Background: The pathogenic bacterium on the hands of healthcare workers (HCWs) is considered as the main route of spread of nosocomial infections. The study was aimed to determine the bacterial contamination of hands of HCWs during routine patient care in a large teaching hospital. Material and Methods: This study includes a total of 200 consecutive, non-repetitive participants. Doctors, medical students, nurses and attendants from the Department of Anaesthesia and Department of Medicine of the hospital were included, who were actively providing health care. The informed consent was taken from all the participants of the study. The hand impressions of the participants were taken on 5% blood agar and processed as per guidelines. After taking the hand impressions, hand cleaning of each participant was done by standard preparation of 70% isopropyl alcohol-based hand rub. The hand impressions from the participants were again taken on 5% blood agar plates. Results: Out of 200 samples collected from HCWs, 95 (47.5%) samples showed growth of microorganisms. Most commonly isolated microorganism was Staphylococcus aureus present in 70% of nurses, 60% of students and 40% of attendants. Prevalence of Acinetobacter spp. was also found to be high among nurses (20%), followed by attendants, sweepers (16%) and doctors (4%). Conclusions: Hands of hospital staff are colonised with pathogenic bacteria, especially Gram-positive such as S. aureus. Hand hygiene is an effective method of reducing the bacterial flora on the hands of the HCWs.

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A 69-year-old male presented with complaints of high-grade intermittent fever for 1 week. The patient is a known case of coronary artery disease. Seven blood samples were sent for culture, and all the samples grew beta haemolytic colonies, which were identified as Streptococcus pluranimalium. The transoesophageal echo finding of the patient was suggestive of aortic ring abscess. Since the patient was not willing for invasive measures, he was conservatively treated with appropriate antibiotics, to which he responded symptomatically.

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Background and Objectives: Liver abscess is a potentially life-threatening condition with a majority of cases being amoebic in tropical countries. In India, poor sanitary conditions and low socioeconomic status contribute to the endemicity of amoebiasis. As treatment modalities are different for amoebic and pyogenic liver abscesses (ALA and PLA, respectively), it is necessary to differentiate them. Facilities for amoebic culture are not available and microscopy is nondiscriminatory, hence serological tests are greatly relied upon for the identification. No studies have been done so far in our region on patients admitted with liver abscess. The aims of this study were to find the aetiology of liver abscess in admitted patients and to know the proportion of amoebic liver abscess using serological techniques. Materials and Methods: The study was done for 6 months and the sample size was 50. The ultrasonography (USG)-guided aspirates were subjected to microscopy and aerobic and anaerobic cultures on appropriate media. Serum samples of all patients were examined for Entamoeba histolytica immunoglobulin G (IgG) antibodies by enzyme-linked immunosorbent assay (ELISA) along with 35 control samples. Results: Of the 50 patients, 34 were diagnosed as amoebic by (ELISA) (68%), 11 as pyogenic (22%) and five as indeterminate (10%). Alcohol was found to have a higher association with ALA than with PLA. Of the 34 amoebic liver abscesses, only four (8.8%) showed motile trophozoites. The mortality rate was only 2.2%. Conclusion: Rapid diagnosis with serology and prompt treatment can result in reduced hospital stay in cases of ALA.

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BACKGROUND AND OBJECTIVE: An outbreak of diphtheria was identified in Malappuram district, North Kerala, India, from July 2016 onwards as a sudden increase in the number of throat swabs with suspected diphtheria received in the Microbiology Lab of Government Medical College Manjeri, Malappuram district. The objective of this study was to confirm the outbreak and identify the organisms and their antibiotic sensitivity patterns. MATERIALS AND METHODS: A total of 554 throat swabs were received from July 2016 to June 2017 from neighbouring areas of Malappuram district for culture and sensitivity. Elek gel precipitation test and polymerase chain reaction of the isolates were performed at the State Public Health Lab, Thiruvananthapuram. All isolates of Corynebacterium diphtheriae possess diphtheria toxin gene. Antibiotic sensitivity tests were performed. Repeat swabs were not included in the study. RESULTS: Of the 29 (5.23%) confirmed cases, 18 (62%) were male and 11 (38%) were female. Fourteen (48.3%) cases were between 6 and 10 years of age with nine (64%) males and five (36%) females. Seven (24.2%) were between 11 and 15 years of age with four (57%) males and three (43%) females. Three (10.3%) were between 16 and 20 years of age (all males). Two (6.9%) cases from 26 to 30 years and one (3.4%) each from 21 to 25 years, 31–35 years and 36–40 years. All isolates were Penicillin sensitive. CONCLUSIONS AND DISCUSSION: Our study identified an upsurge of diphtheria in Malappuram district, North Kerala, among older children and adolescents. Most of them are partially immunised and immunity to diphtheria declines over time. Adults and adolescents thus become susceptible later in life. This emphasises the need for booster vaccination above five years in the endemic areas and making antidiphtheritic serum available in all major hospitals of these regions.

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INTRODUCTION: Urinary tract infections (UTIs) are among the most common bacterial infections. Escherichia coli remains the most frequent cause of UTIs. More important is the increase in resistance to some antimicrobial agents. Furthermore, bacterial species are capable of living in a biofilm. There is increasing evidence for the role of bacterial biofilm in causing recurrent UTIs. AIM: The present study aimed to evaluate the ability of E. coli, isolated from UTIs to form a biofilm, and its association with catheterisation and to correlate the role of biofilms with their antimicrobial resistance. MATERIALS AND METHODS: A total of 403 urine samples were processed. All the isolated E. coli strains (226) were grown in Luria broth and were incubated overnight in high-glucose Dulbecco's modified Eagle's medium using a microtitre plate. The plate was stained with crystal violet, and the biofilm was quantified using an enzyme-linked immunosorbent assay plate reader at 570 nm. An optical density value more than that of the mean negative control plus three standard deviations is taken as positive for biofilm production. The antibiogram was done using the Kirby–Bauer disk diffusion method. RESULTS AND DISCUSSION: Of 226 strains, 54.4% were found to produce biofilms. Of them, 81.3% of patients were catheterised. Most of them were found to be resistant to commonly used antibiotics such as Cephalosporins, Quinolones and Aminoglycosides. Imipenem and Nitrofurantoin are the most effective antibacterial agents, showing 77.3% and 73.2% sensitivity, respectively. CONCLUSION: The biofilm assay using a microtitre plate is convenient and useful in screening the biofilm producers. Catheterisation is a risk factor for biofilm production, and catheter care is of paramount importance to prevent catheter-associated UTI.

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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.

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Colistin is an important antibiotic against multidrug-resistant (MDR) Gram-negative bacteria (GNB), particularly Escherichia coli, Klebsiella pneumoniae, Pseudomonas aeruginosa and Acinetobacter baumannii. Although it is an old antibiotic, Colistin has emerged as an important agent in the treatment of GNB infections, especially those caused by MDR pathogens in hospitalised patients. Limited therapeutic option against Carbapenem-resistant Gram-negative bacilli (E. coli, Klebsiella, Pseudomonas and Acinetobacter) is one of the major problems in clinical practice. The current review discusses the pharmacology, toxicity, drug resistance, importance of therapeutic drug level monitoring and cost of one of the most important antibiotics in the current practice.

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Melioidosis is a disease of public health importance in South East Asia and northern Australia, caused by a Gram negative soil-dwelling bacterium, Burkholderia pseudomallei. Sporadic cases are reported from other parts of the world, including India, while detailed information is unavailable. The disease is mainly seen in the population who are in prolonged contact with soil and are diabetic. There is a close association of the disease with rainfall. The primary manifestation is pneumonia with or without septicaemia, but other organs like bone and joints, liver, spleen, lymph nodes may also be affected. Despite improvements in antibiotic therapy, melioidosis is still associated with a significant mortality attributable to severe sepsis and its complications. The accurate and practical diagnostic set up is still unavailable in many parts of India, which leads to under- or mis-diagnosis, and fatality. The disease needs more attention from the microbiologists, clinicians and researchers from India, since studies related to the pathogenesis and virulence factors, preventive measures, vaccine prospects, and better management may reduce the burden of the disease.

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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.

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Virus-associated hemophagocytic syndrome (VAHS) is a rare complication in early cytomegalovirus (CMV) infection. It is a life-threatening condition characterized by prolonged fever, hepatosplenomegaly, and cytopenia. There is no standard therapy for VAHS and the clinical course is variable. Herein, we report a 3-month-old boy whose clinical and laboratory findings were consistent with CMV infection-associated hemophagocytic syndrome. In spite of prompt diagnosis and treatment, the infant expired due to progressive respiratory failure.

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Exophiala jeanselmei is clinically redefined as a rare agent of subcutaneous lesions of traumatic origin eventually causing mycetoma. A case of eumycetoma of foot in a 42-year-old male was clinically diagnosed as dermoid cyst and was microbiologically confirmed as eumycetoma. This case is reported for its uncommon clinical presentation and etiological agent, Exophiala jeanselmei. The patient recovered completely after treatment with Ketoconazole.

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Aspergillosis is one of the most common fungal infections in humans caused by Aspergillus species. However, not all Aspergillus species possess the same antifungal susceptibility patterns. Aspergillus terreus is an emerging pathogen which commonly occurs in infections of immunocompromised individuals and is associated with high mortality. It gains its importance in being mostly resistant to treatment with amphotericin B. The four cases of A. terreus reported here highlight the fact that fungal infections caused by this species are on the rise. Early diagnosis and prompt treatment are important to prevent an unfavorable outcome. The optimal therapy for infections caused by this emerging pathogen is still to be ascertained.

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Enterobacteriaceae producing both Amp C beta lactamases and extended-spectrum beta lactamases (ESBLs) have been increasingly reported worldwide. While the phenotypic tests for ESBL is standardised and used widely, it is not so for Amp C. When they coexist they may mask each other's detection phenotypically. We undertook this study to detect the concurrent occurrence of Cefotaxime (CTX)-M and plasmid Amp C in clinical isolates of Enterobacteriaceae by phenotypic and genotypic methods. One hundred clinically significant isolates of Escherichia coli (E. coli; 43), Klebsiella pneumoniae (K. pneumoniae; 43) and Proteus mirabilis (P. mirabilis; 14) were included in the study. Antibiotic susceptibility testing to various classes of antimicrobials was performed by disc diffusion using Clinical Laboratory Standards Institute (CLSI) guidelines. Isolates were screened for production of ESBL by CLSI method and Amp C beta lactamase by inhibitor based method using boronic acid and cloxacillin. Polymerase chain reaction (PCR) was performed for the detection of plasmid Amp C genes and bla_CTX-M . Plasmid Amp C genes were detected in 27 isolates which included CIT (Origin Citrobacter freundii): 14; DHA (Dhahran Hospital in Saudi Arabia): 12; EBC (Origin Enterobacter cloacae): 1. Bla_CTX-M was detected in 51 isolates. Both coexisted in one E. coli and two K. pneumoniae. In one of the K. pneumoniae isolate, all phenotypic tests employed were negative. A high degree of cross resistance to other classes of antimicrobials was observed. Carbapenem resistance was noted in 21 isolates. The concurrent occurrence of Amp C and CTX-M is not common in clinical isolates of Enterobacteriaceae. Phenotypic tests perform poorly when these enzymes are coproduced.

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Introduction : Non-Typhoidal Salmonella (NTS) species are important food borne pathogens with acute gastroenteritis being the most common clinical manifestation. Certain serovars of Salmonella show a much higher predilection for causing bacteraemia. Though NTS infections are associated with significant morbidity and mortality worldwide; data on NTS in India are limited. Objectives : The aim of this study was to describe the prevalence and epidemiology of Non-Typhoidal Salmonella (NTS) in central Kerala, and to compare their sensitivity patterns with typhoidal Salmonella. Methods : A retrospective study was done on the blood and stool culture samples received by the microbiology laboratory of a tertiary care centre from January 2012 to August 2014. NTS were isolated and identified using standard bacteriological methods, including serotyping with specific antisera. The geographical distribution, co-morbid conditions and antibiotic sensitivity patterns were analyzed. Results : A total of 15 cases of NTS were isolated of which three isolates were associated with bactaeremia. Most common isolates were S. Typhimurium and S. Enteritidis. We also isolated S. Weltreveden from two patients. Conclusion : Knowledge of the prevalent of serotypes and their sensitivities would be useful for epidemiological and treatment purposes.

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Histoplasmosis is commonly found in US and central America; recently cases were diagnosed from Kerala also. The causative organism Histoplasma capsulatum is found abundant in soil mixed with bird or bat guano. Infection occurs by the inhalation of small oval conidia which can enter the terminal bronchioles and then the alveolar spaces. They multiply inside the macrophages and the cellular immune system of the host decides the outcome. Usually it is a self limiting pulmonary infection, but it can vary from mild pneumonitis to severe acute respiratory distress syndrome. If primary manifestation progresses, disseminated histoplasmosis involving liver, spleen, bone marrow, adrenal gland and mucocutaneous membranes result. Skin test antigen is used in epidemiological studies to find the true extent of infection. Rapid diagnosis is possible with histoplasma antigen detection but serology is useful only in certain cases. Histopathology aids the diagnosis a lot. But the fungal culture remains the gold standard for the confirmation. Histoplasmosis is successfully treated with azoles if it is a mild infection and if severe liposomal Amphotericin B is used initially and then switched over to Itraconazole to be continued for several months.

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