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

Se aislaron 110 cepas que pertenecían a siete especies de Candida procedentes de pacientes diabéticos con distintas formas de candidiasis, encontrándose 53 aislamientos de Candida albicans (47%), 36 de Candida tropicalis (33%), 9 de Candida glabrata (8%), 4 de Candida parapsilosis (4%), 2 de Candida guilliermondii (2%), 5 de Candida krusei (5%) y 1 de Candida kefyr (1%). En las 53 cepas de C. albicans aisladas se estudió la expresión de factores de virulencia tales como la hidrofobicidad de la superficie celular (CSH), adherencia a células epiteliales bucales humanas (BEC) y actividad enzimática. La actividad proteolítica se detectó en el 100% de las cepas de C. albicans, mientras que la producción de fosfolipasa se detectó en 52 cepas (98%). Se estudió la variación fenotípica y su influencia en factores de patogenicidad en dos cepas de C. albicans, procedentes de boca y vagina respectivamente, y en la cepa patrón C. albicans NCPF 3153A. Se les indujo la variación fenotípica mediante exposición a luz UV y se valoró el grado de expresión de los factores de virulencia por las diversas formas morfológicas obtenidas. Se obtuvieron tres variaciones morfológicas de C. albicans: forma de estrella (S), rugosa (W) y anular (R), a partir de la variedad lisa original (O). La actividad proteinasa fue mayor en el tipo W, seguida por el tipo R, y por el tipo O; el tipo S fue el de menor actividad proteolítica. La actividad fosfolipasa fue mayor en el tipo O, seguida por el tipo R; los tipos W y S presentaron una actividad fosfolipasa menor. La expresión de la CSH y de la adherencia fue superior en el tipo O, seguida por el tipo R y el tipo W, y finalmente el tipo S. Las variaciones fenotípicas de C. albicans presentan una expresión diferenciada de factores de virulencia y ello puede proveer a un tipo morfológico particular de ciertas ventajas, facilitando el inicio de una candidiasis(AU)

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(AU9

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.

Slime production a virulence marker in Pseudomonas aeruginosa strains isolated from clinical and environmental specimens: a comparative study of two methods.

Detection of slime in Pseudomonas aeruginosa can be useful in understanding the virulence of this organism. Here, comparative studies of two phenotypic methods using the tube method and the spectrophotometric method for slime production from 100 clinically and 21 environmentally significant isolates of P. aeruginosa were performed. A total of 68 isolates were positive by either of the tests whereas only 34 were positive by both the tests. The tube method detected slime significantly in more number of isolates than the spectrophotometric method. The tube test was found to be superior to the spectrophotometric method in ease of performance, interpretation and sensitivity. Among the clinical isolates, systemic isolates produce less slime compared to wound, respiratory and urinary isolates. Isolates from the hospital environment produced more slime indicating that this virulence marker helps the organism to survive for longer periods and cause nosocomial infections.

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.

Phenotypic switching and its influence on expression of virulence factors by Candida albicans causing candidiasis in human immunodeficiency virus-infected patients.

PURPOSE: The purpose of the present study was to determine the degree of expression of virulence factors such as adherence, cell surface hydrophobicity (CSH) and production of proteinase by different morphological forms of Candida albicans causing oral candidiasis in human immunodeficiency virus (HIV)-infected individuals. METHODS: C. albicans 3153A and two strains isolated from oral thrush in HIV infected individuals 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 studied. RESULTS: 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 production was greatest with the W type followed by the R type and O type. The S type produced the least proteinase. Expression of cell surface hydrophobicity and adherence was greatest in the O type followed by the R and then the W type and finally the S type. CONCLUSIONS: The 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.

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.

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.

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.

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.

Student attitudes towards communication skills training in a medical college in Western Nepal.

CONTEXT: Previous studies have shown that students have both positive and negative attitudes towards communication skills training. However, studies in Nepal are lacking. OBJECTIVES: The present study was carried out to determine the positive and negative attitudes of student respondents using the previously validated communication skills attitude scale (CSAS) (see Appendix). METHODS: The study was carried out among third- and fourth-semester students at the Manipal College of Medical Sciences, Pokhara, Nepal. These students are in the pre-clinical part of their course and learn the basic science subjects through an integrated, system-based curriculum. Gender, age, nationality of the respondents, occupation of parents, medium of instruction at school, attitude towards communication skills training during the clinical years, and self-rating of communication abilities were recorded. Association of the positive and negative attitudes with these variables was determined using appropriate statistical tests (p < 0.05). FINDINGS: A total of 123 students participated in the study; 74 (60.2%) were male, and 104 (84.5%) had studied in English-medium schools. The median positive attitude score was 51 (inter-quartile range 7). Nationality and attitude towards communication skills training during the clinical years showed a significant association. The mean negative attitude scale score was 31.18 (SD = 4.96). A significant association was noted with attitude towards communication skills training during the clinical years. Both scales range from 13-65, with higher scores indicating stronger attitudes. CONCLUSIONS: Communication skills training should be modified and strengthened. Formal courses during the clinical years are required. Training sessions for the faculty and further studies across different semesters and in different medical colleges in Nepal are needed.

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.

Prevalence of Moraxella catarrhalis infections of the lower respiratory tract in elderly patients.

OBJECTIVES: To determine: *rate of isolation of Moraxella catarrhalis in elderly patients having lower respiratory tract infection *The antibiotic susceptibility pattern of Moraxella catarrhalis isolates. *The predisposing factors responsible for Moraxella catarrhalis infections of lower respiratory tract. MATERIAL AND METHOD: This is a prospective study carried out at Manipal Teaching Hospital, Pokhara during May 2002 to April 2003. A total of 716 sputum samples collected from patients with suspected lower respiratory tract infection were investigated. The samples were subjected to Gram stain, culture and sensitivity. Sputum samples were inoculated on blood and chocolate agar and incubated at 37 degrees C for 18 to 24 hours in candle jar containing 3-5% Co2. For the identification of bacterial isolates, the cultural and colonial characteristics were recorded and identified with the use of biochemical test. The susceptibility of bacterial isolates to antimicrobial agent was performed following Kirby-Bauer method. RESULT: Out of the 716 samples, 355 (49.58%) grew normal commensals of the upper respiratory tract. Respiratory pathogens were recovered from 361 (50.41%) samples. The most common respiratory pathogen was H. influenzae (26. 86%), followed by S. pneumoniae (21.16%). Moraxella catarrhalis accounted for 6.90%. Rate of isolation of Moraxella catarrhalis was higher in males than in females. Out of 25 isolates 16 were from males and 9 were from females. Frequency of isolation of Moraxella catarrhalis was more frequently seen in age group 61-70 years. It was most commonly recovered in winter. The most effective antibiotics were amoxicillin-clavulanate and ceftriaxone and least was ampicillin. CONCLUSION: Moraxella catarrhalis isolation from sputum especially in persons above 60 years of age and in the absence of other well established pathogens should not be disregarded as they can cause lower respiratory tract infection in these individuals.

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.

Effect of trace elements on surface hydrophobicity and adherence of Escherichia coli to uroepithelial cells.

Trace elements have significant effect on the physiology of bacteria. Variation in the concentration of trace elements may affect the expression of virulence by microorganisms. The effect of trace elements on hydrophobicity and adherence of E.coli to uroepithelial cells was studied. Increasing concentrations of Ca2+, Mg2+, Fe3+ and Zn2+ significantly decreased the surface hydrophobicity. Toxic trace elements like Co2+, Cu2+, Mn2+ and Ni2+ did not alter surface hydrophobicity. With regards to adherence of E.coli to uroepithelial cells, only Mg2+ had significant effect. Toxic trace elements decreased the rate of cell adherence. The pathogenic strains of E.coli showed higher surface hydrophobicity and better cell adherence compared to the nonpathogenic strains. There was good correlation between surface hydrophobicity and cell adherence at higher concentrations (0.1 to 0.2mM) of Fe2+ and Zn2+. The results indicated that trace elements can significantly affect surface hydrophobicity and adherence of E.coli to uroepithelial cells. Such effect may have a significant impact on the initial stages of bacterial infection.

Spectrum of opportunistic infections in AIDS cases.

Human Immunodeficiency viruses are the initial causative agents in AIDS, but most of the morbidity and mortality in AIDS cases result from opportunistic infections, Identification of such pathogen is very important for clinicians and health planners to tackle the AIDS epidemic in more effective manner. The present study describes the clinical and laboratory profile of 100 AIDS causes who presented to a referral hospital. Oral candidiasis (59.00%) was found to be the most common opportunistic infection, followed by tuberculosis (56.00%), Cryptosporidium infection (47.00%) and Pneumocystis carinii (7.00%). Presence of oral candidiasis and weight loss is highly predictive of low DC4 count and can be considered as a marker of HIV disease progression. The patients coinfected with HIV and tuberculosis are also on rise. Recognition of dual infection and taking adequate steps to deal with this epidemic is needed. As Cryptosporidium infection was detected in large number, provision of safe drinking water and maintaining good hygiene is important for prevention. Early diagnosis of opportunistic infection and prompt treatment, delays the progression towards AIDS. 91.00% of patients were infected with HIV1 and 4.00% had HIV2 infection and 5.00% were dully infected. 87.00% of patients were males and 13.00% were belonging to 21-40 years of age. Majority of them were belonging to lower socioeconomic status and heterosexual route of transmission was the commonest mode of spread.