Streptococcus pneumoniae infections in western Nepal.

We conducted a study to determine the prevalence of antibiotic resistance among clinical isolates of S. pneumoniae. This study was conducted from January 2000 to August 2007 at a tertiary care teaching hospital in Nepal. The isolates were identified based on standard bacteriological techniques. Antibiotic susceptibility testing used the Kirby-Bauer disc diffusion method; penicillin resistance was confirmed by agar dilution method. During the study period, there were 312 S. pneumoniae isolates. Penicillin, trimethoprim-sulfamethoxazole, erythromycin, tetracycline and chloramphenicol resistance were observed in 5, 34.3, 7.4, 11.1 and 0.4% of isolates, respectively. Resistance to all tested antibiotics declined with time except for penicillin, in which resistance increased. Penicillin-resistant S. pneumoniae were significantly co-resistant to erythromycin. Co-resistance to tetracycline and erythromycin were observed in trimethoprim-sulfamethoxazole resistant isolates. Penicillin resistance is increasing; therefore, measures to ensure judicious use of beta-lactams and macrolides (inducers of penicillin resistance) should be advocated to control the development of penicillin-resistant S. pneumoniae.

Effect of phenotypic switching on expression of virulence factors by Candida albicans causing candidiasis in diabetic patients.

A total of 110 strains belonging to seven species of Candida were isolated from various forms of candidiasis in diabetic patients. They were Candida albicans 53 (47%), Candida tropicalis 36 (33%), Candida glabrata 9 (8%), Candida parapsilosis 4 (4%), Candida guilliermondii 2 (2%), Candida krusei 5 (5%) and Candida kefyr 1 (1%). All 53 strains of C. albicans isolated were observed to express virulence factors such as cell surface hydrophobicity (CSH), adherence to human buccal epithelial cell (BEC) and proteinase activity (100%), while phospholipase activity was observed in 52 (98%). Phenotypic switching and its influence on the pathogenicity of C. albicans were studied. Two C. albicans strains isolated from oral and vaginal thrush, respectively, in diabetic individuals, and the control strain C. albicans NCPF 3153A were induced to undergo phenotypic switching by exposure to UV light and the degree of expression of virulence factors by the different morphological forms was determined. Three different morphological forms of C. albicans were obtained, namely Star (S), Wrinkled (W) and Ring (R) types from the original Smooth (O) variety. It was found that proteinase activity was greatest with the W type followed by the R type then the O type. The S type produced the least proteinase. The phospholipase activity was greatest with O type followed by R type. The W and S types produced the least phospholipase. Expression of CSH and adherence was greatest in the O type followed by the R and then the W type and finally the S type. Differential expression of virulence factors occurs with different phenotypic forms of C. albicans and this may provide a particular morphological type with a distinct advantage over other types in causing candidiasis.

A 6 year Geohelminth infection profile of children at high altitude in Western Nepal.

BACKGROUND: Geohelminth infections are a major problem of children from the developing countries. Children with these infections suffer from developmental impairments and other serious illnesses. This study aimed to measure the prevalence of geohelminth infection, infection intensity as well as the change in the intensity in children from Western Nepal over years. METHODS: This 6-year hospital based prospective study at the Manipal Teaching Hospital, Pokhara included children (< 15 years) visiting the hospital from Kaski and 7 surrounding districts. Samples were also collected from children in the community from different medical camps. Three stool samples from every child were processed using direct and concentration methods. The Kato-Katz technique was used for measuring the intensity of infection. RESULTS: The overall prevalence in hospital - attending children was 9.2% with 7.6% in preschool (0 - 5 y) and 11.0% in school-age (6 - 15 y) children, and in community 17.7% with 14.8% in pre-school and 20.5% in school-age children. Ascaris lumbricoides, Trichuris trichiura, Ancylostoma deodenale and Strongyloides stercoralis were the common geohelminths with a gradual decrease in worm load over the years. School-age children were found to be significantly more prone to geohelminth infection as compared to preschool children, but no statistical difference was detected by gender, district as well as season. CONCLUSION: This heavy infection of geohelminths in children should be corrected by appropriate medication and maintaining strict personal hygiene. Health education, clean water, good sewage management and a congenial environment should be ensured to minimise infection.

Intestinal protozoal infestation profile in persistent diarrhea in children below age 5 years in western Nepal.

Unlike acute diarrhea, the role of pathogens in persistent diarrhea in children in Nepal is unclear. Protozoal parasites are suspected to be a major cause. The study was carried out to find the association between protozoal agents and persistent diarrhea in children below age 5 years from western Nepal. Stool samples were collected from 253 children with persistent diarrhea, from 155 children with acute diarrhea (disease controls) and from 100 healthy children from the community (normal controls). Of 253 children with persistent diarrhea, 90 (35.5%) had protozoal infections, 63 (24.9%) helminthic infections, 32 (12.6%) had bacterial infections and 16 had mixed infections. Giardia lamblia was the most prevalent (67.7%), followed by Entamaeba histolytica (27.7%). HIV infection and severe malnutrition were associated with Cyclospora cayetanensis and Cryptosporidium spp causing persistent diarrhea. We conclude that stool microscopy should be routinely performed in children with persistent diarrhea since protozoal infections can be cured with effective treatment and control can be achieved by proper health education.

Paraffin baiting system for demonstration of growth and biofilm production in Pseudomonas aeruginosa.

Pseudomonas aeruginosa is one of the commonest pathogens among the pseudomonads. This organism can grow in minimal nutritional requirements. Because of the ability of pseudomonads to grow on paraffin is not commonly found among other human pathogens and the primary human pathogen being P. aeruginosa, we studied the adaptation of this organism to paraffin baiting system for growth and biofilm formation. Strains were tested for the capacity to use paraffin as the sole source of carbon using Czapek's minimal salt medium. Of the 53 clinical isolates of P. aeruginosa, 20 strains exhibited growth by 24 hrs and 42 strains by 48 hrs. The remaining strains did not show any growth in the paraffin baiting system. The oxidase test with the paraffin baiting system was also performed. This simple and inexpensive method can be used to isolate and demonstrate the biochemical and biofilm forming capacity of the organism.

Induction of resistance to fluoroquinolones in clinical and environmental isolates of Pseudomonas aeruginosa.

An attempt to induce resistance to ciprofloxacin in vitro was made against clinical and environmental isolates of Pseudomonas aeruginosa. This in vitro manipulation of strains resulted in the increase of minimum inhibitory concentration from 0.4 microg/ml to 1 microg/ml of the original strains to 9.0 to 12.5 microg/ml indicating development of resistance to ciprofloxacin and a major decrease in the size of zone diameters of norfloxacin and ofloxacin indicating cross resistance to these agents. Results indicate the induced resistance to ciprofloxacin can promote development of cross resistance to other fluoroquinolones. This suggests that caution should be taken while using fluoroquinolones for the treatment of Pseudomonas aeruginosa infections.

Isolation & antimicrobial susceptibility of Shigella from patients with acute gastroenteritis in Western Nepal.

BACKGROUND & OBJECTIVE: Shigellae play an important role as a causative organism of acute gastroenteritis, which is a global health problem with significant morbidity and mortality in especially in developing countries. This study was carried out to determine the isolation and pattern of antimicrobial resistance of Shigella in patients with acute gastroenteritis in western Nepal. METHODS: The study included all patients with acute gastroenteritis who visited a tertiary care hospital at Pokhara, Nepal during a 2-year period (2002-2004). The isolates was confirmed as Shigella by biochemical reaction and slide agglutination test using specific antisera. Antibiotic sensitivity test was determined by agar diffusion method and minimum inhibitory concentration (MIC) of the drugs was detected. RESULTS: Of the 770 stool samples, 83 (10.8%) yielded Shigella. Shigella flexneri caused 56 (67.4%) of the total cases of shigellosis followed by S. dysenteriae 12 (14.5%), S. sonnei 10 (12%) and S. boydii 5 (6%). Of the 83 isolates, 67 (80.7%) showed resistance to various drugs and 62 (74.7%) were resistant to two or more drugs. Resistance to cotrimoxazole was 80.7 per cent followed by tetracycline 74.7 per cent, ampicillin 53.0 per cent, nalidixic acid 31.3 per cent and ciprofloxacin 2.4 per cent. The MIC(50) and MIC(90) values of those drugs were also very high. All isolates were sensitive to cefotaxime and ceftriaxone. INTERPRETATION & CONCLUSION: The findings of our study suggested that Shigellae was an important etiological agent for acute gastroenteritis, with a high rate of drug resistance and requires constant monitoring in this region.

Frequency and susceptibility profile of pathogens causing urinary tract infections at a tertiary care hospital in western Nepal.

INTRODUCTION: Urinary tract infection (UTI) is the most common nosocomial infection among hospitalised patients. Area-specific monitoring studies aimed to gain knowledge about the type of pathogens responsible for UTIs and their resistance patterns may help the clinician to choose the correct empirical treatment. Recent reports have shown increasing resistance to commonly-used antibiotics. We aimed to study the antibiotic resistance pattern of the urinary pathogens isolated from hospitalised patients. METHODS: Three urine samples were collected by the mid-stream "clean catch" method from 1,680 clinically-suspected cases of urinary tract infections from inpatients of various clinical departments during one year. The samples were tested microbiologically by standard procedures. Antibiotic susceptibility of the isolated pathogens was tested for commonly-used antibiotics by Kirby-Bauer technique according to NCCLS guidelines. RESULTS: Significant bacteriuria was present in 71.7 percent of the samples, 17 percent were sterile, 4.8 percent showed insignificant bacteriuria, and 6.5 percent non-pathogenic bacteriuria. The most common pathogens isolated were Escherichia coli (59.4 percent), Klebsiella spp (15.7 percent) and Enterococcus faecalis (8.1 percent). The mean susceptibility was high for amikacin (87.2 percent), ciprofloxacin (74.8 percent), ceftazidime (71.5 percent) and gentamicin (70.4 percent) but low for nitrofurantoin (35 percent), cephalexin (49.7 percent) and ampicillin (50.5 percent). Escherichia coli was found to be most susceptible to amikacin (98 percent) followed by gentamicin (87.9 percent), ceftazidime (80.8 percent), norfloxacin (78.4 percent) and cotrimoxazole (77.9 percent). CONCLUSION: A high isolation rate of pathogens from urine samples of clinically-suspected UTI shows a good correlation between clinical findings and microbiological methods. The antibiotics commonly used in UTIs are less effective. Since the present study was a cross-sectional study, regular monitoring is required to establish reliable information about resistance pattern of urinary pathogens for optimal empirical therapy of patients with nosocomial UTIs.

Characterization of Candida species isolated from cases of lower respiratory tract infection.

OBJECTIVES: (1) To identify and characterize the Candida species isolates from lower respiratory tract infection. (2) to determine the rate of isolation of Candida species from sputum samples. METHODS: This study was carried out in the Department of Microbiology, Manipal Teaching Hospital, Pokhara, Nepal from June 2002 to January 2003. A total of 462 sputum samples were collected from patients suspected lower respiratory tract infection. The samples were processed as Gram staining to find out the suitability of the specimen, cultured on Sabouraud's Dextrose Agar (SDA) and also on blood agar and chocolate agar to identify the potential lower respiratory tract pathogens. For the identification of Candida, sputum samples were processed for Gram stain, culture, germ tube test, production of chlamydospore, sugar fermentation and assimilation test. For the identification of bacteria, Gram stain, culture, and biochemical tests were performed by standardized procedure. RESULT: Out of 462 samples, 246 (53.24%) samples grew potential pathogens of lower respiratory tract. Among them Haemophilus influenzae 61(24.79%) and Streptococcus pneumoniae 57 (23.17%) were the predominant bacterial pathogens. Candida species were isolated from 30 samples (12.2%). The majority of Candida species amongst the Candida isolates were Candida albicans 21(70%) followed by Candida tropicalis 4(13.33%). Candida krusei 3(10%), Candida parapsilosis 1(3.33%) and Candida stellatoidea 1(3.33%). The highest rate of isolation of Candida was between the age of 71 and 80. CONCLUSION: Candida isolation from sputum samples is important as found in the present study in which Candida species were the third most common pathogen isolated from patients with lower respiratory tract infection.

Effect of "HAART" on CD4+ T lymphocyte counts in HIV seropositive south-Indian individuals--a follow up study.

Because of the drastic cut down in the prices of antiretroviral drugs, the antiretroviral therapy has become more affordable in India. 16 HIV-1 infected individuals who were on HAART were followed up for 12 months. The CD4 + T Cell count estimation was done at mean intervals of 6 months and 12 months. Antiretroviral therapy showed good response in 14(87.5%) and deterioration in 2 (12.5%) cases.

Emergence of multidrug-resistant Salmonella typhi in rural southern India.

Multidrug-resistant Salmonella typhi has spread to many parts of India, causing severe therapeutic problems. Of the 305 clinically suspected cases of enteric fever seen at Kasturba Hospital in Manipal, Karnataka between January 1990 and June 1991, Salmonella bacteremia was detected in 134 patients; 102 of these were caused by S. typhi. Eighty (78.4%) of the isolates from S. typhi-infected patients were resistant to conventional antibiotics used in the treatment of typhoid fever (i.e., ampicillin, chloramphenicol, and trimethoprim-sulfamethoxazole). No in vitro resistance was observed to ciprofloxacin, norfloxacin, and gentamicin. Major complications were seen in four patients infected by multiresistant S. typhi; three responded well to ciprofloxacin. The fourth patient died of hemorrhage and acute renal failure, even after receiving ciprofloxacin. In addition, less severe complications such as hepatitis and jaundice were observed in 12 other patients. Seventy-six of the multiresistant S. typhi belonged to phage type O biotype II and four were of untypeable Vi strains. The emergence of multidrug-resistant S. typhi has necessitated the use of fluoroquinolones in the therapy for enteric fever.

In vitro adhesiveness of Bacteroides fragilis group in relation to encapsulation.

The adhesive properties of the B. fragilis group to various erythrocytes were studied by haemagglutination technique and compared to encapsulation. Of the 61 strains of Bacteroides groups tested, only 40 strains of B. fragilis exhibited haemagglutination. Treatment of the organisms with carbohydrates and trypsin had no effect on haemagglutination, but boiling for one hour completely destroyed this property. Haemagglutinating activity of B. fragilis was found to be the highest with chick and tanned sheep erythrocytes. The presence of capsule was observed in 46 strains of B. fragilis group. Of the 40 strains of B. fragilis which exhibited agglutination, capsule was found only in 37. The findings of the study suggest a polysaccharide nature for the adhesin. The different patterns of haemagglutination, suggest the involvement of more than one adhesin. Capsule in capsulated strains and fimbriae in non-capsulated strains might be mediating the property of haemagglutination.

Correlation of phospholipase A & enterotoxin production by Salmonella typhimurium with reference to virulence parameters.

Seventy strains of S. typhimurium along with 10 strains each of S. typhi and S. paratyphi B, 3 strains of S. enteritidis and 2 strains of S. senftenberg were examined for phospholipase A (PLA) activity. S. typhimurium positive for PLA activity (17.14%) were subjected to study of other virulence parameters like enterotoxin production, haemolytic activity, surface hydrophobicity and antibiotic sensitivity. A high degree of correlation was observed between PLA activity and enterotoxin production, antibiotic sensitivity and cell surface hydrophobicity respectively.

Copper resistance & its correlation to multiple drug resistance in Salmonella typhi isolates from south Karnataka.

A study was conducted to ascertain the prevalence of copper resistant Salmonella typhi isolates in south Karnataka. Of the 186 strains studied, 26 (13.97%) were found to be copper resistant. Among the copper resistant strains 19 (73.08%) were found multi drug resistant. All copper resistant strains remained uniformly sensitive to ceftriaxone, ciprofloxacin and norfloxacin. Multiple drug resistance was exclusively associated with E1 phage types.

Acinetobacter baumannii--an emerging nosocomial pathogen in the burns unit Manipal, India.

A retrospective study on 113 patients admitted to the burns unit over a period of 12 months was undertaken to investigate the emerging trend of Acinetobacter baumannii infection. Wound infection with A. baumannii was seen in 13 (11.5%) patients. Of these six (46%) patients developed septicaemia with the same bacteria and five (38%) expired. The virulence of these bacteria, drug susceptibility pattern and its role as a nosocomial pathogen is discussed.

Nosocomial complicated urinary tract infections: A rural based referral hospital study.

A three-year retrospective study of urinary tract infection in 891 patients was conducted at a rural based hospital in coastal Karnataka, South India. Eighty-eight per cent of the patients had complicated urinary tract infections with predominantly nosocomial organisms such as Klebsiella as opposed to the normally more common uncomplicated infection with E. coli. The reasons for this seem to be the delay in referral, resulting in late correction of the underlying structural causes of the infections. In our series of patients, the association of prolonged indwelling catheter drainage with complicated urinary tract infection suggests that non-urologists should adopt alternative measures such as clean intermittent catheterization, and reduce the period of indwelling catheters to the shortest time possible. More judicious selection and timing of antibiotic therapy might prevent the development of resistant organisms.

Changing anaerobic spectrum in suppurative lung disease: a case report.

A spectrum of three different anaerobes were isolated from a debilitated patient with suppurative lung disease, within a two-year period. Repeated isolation from three consecutive samples and symptomatic relief with metronidazole provide clinical evidence of anaerobic lung infection. This case emphasizes the importance of anaerobic culture in cases of protracted pulmonary suppurative disease.

Mycotic keratitis: a study in coastal Karnataka.

Fungi were isolated from 67 cases out of the 295 cases of corneal ulcers investigated. Aspergillus species and species of Candida were the major fungal members isolated. Allescheria boydii was isolated from 3 cases, having no previous history of injury to the eye or infection with bacterial or viral agents. A boydii corneal infection is a rare occurrence. Higher incidence of mystic keratitis was seen among females than males. No relationship to seasonal changes could be established. Bacterial infection was associated in 46. 27% of the cases of mycotic keratitis and Staphylococcus was the predominant bacterial pathogen observed.

Acanthamoeba keratitis--a report of two cases.

Two cases of unilateral corneal ulcers caused by Acanthamoeba are reported. Neither of the patients had contact lenses at any time. The diagnosis was confirmed by Giemsa stain and cultures of the corneal scrapings.

Cryptosporidium and isospora belli diarrhoea in immunocompromised hosts.

Cryptosporidium and isospora, two of the intestinal coccidian parasites known to be the causative agents of acute diarrhoea in animals, have now emerged as one of the main causes of prolonged life threatening diarrhoea in immunocompromised patients particularly so in patients with AIDS. Between June 1996 and December 1997, a total of 75 immunocompromised patients presenting with diarrhoea were investigated both for Cryptosporidium and Isospora. The study group consisted of cancer and AIDS patients with history of diarrhoea. Cryptosporidium oocysts were detected in 35 patients (46.7%). One of the faecal samples from an AIDS patient with diarrhoea showed the presence of both Cryptosporidium and Isospora oocysts. To the best of our knowledge, this is the second documented report of Isospora associated diarrhoea in an AIDS patient from India. The various techniques used for demonstration of these parasites were modified acid fast staining, Safranine Methylene-blue staining and direct immunofluorescence test.