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Studies on pathogenic dematiaceous fungi. 1. Isolation from natural sources.

Two hundred and twenty-six samples of woody materials, vegetable matter and soil were processed by the direct plating and mouse inoculation technique for the isolation of pathogenic dematiaceous fungi. The species of fungi isolated were Fonsecaea pedrosoi - 13, Cladosporium carrionii - 7 and Phialophora verrucosa -4 isolates. The mouse inoculation technique was found to be much better than direct plating for the recovery of these fungi. Woody plant materials proved to be a good sample source for pathogenic dematiaceous fungi contributing about 90% of the isolates. All the isolates were pathogenic for mice as evidenced by the presence of dark nodular lesions containing fungal elements in the organs of experimentally infected animals.

Animals↗

[A clinical study of respiratory infection isolating non-pathogenic Neisseria by transtracheal aspiration].

Neisseria species other than N. meningitidis and N. gonorrhoeae are generally regarded as commensal bacterial flora of the oropharynx, and little is known regarding cases of these non-pathogenic Neisseria species in the lower respiratory tract. We clinically examined respiratory tract infections from which non-pathogenic Neisseria species were isolated by transtracheal aspiration (TTA). The incidence of non-pathogenic Neisseria isolated was 54 (15.7%) out of 344 episodes of respiratory tract infections with isolated microorganisms from TTA, and was 17.6%, 15.8%, 14.3% for pneumonia, acute bronchitis, and chronic lower respiratory tract infection, respectively. All 54 episodes were isolated with other microorganisms such as alpha-Streptococcus spp. (75.9%), Haemophilus influenzae (25.9%) and anaerobics (22.2%). The isolation ratio according to the age group increased at 45 years of age or more, but did not increase with the advance of age. Predisposing factors were identified such as overt aspiration, iatrogenic procedure and heavy smoking. Cases without overt aspiration that had fevers of 38 degrees C or more or hypoxemia of less than PaO2 70 torr when detecting non-pathogenic Neisseria were observed more frequently in the aged than the non-aged. The findings suggest the detection of non-pathogenic Neisseria by TTA is influenced by the host state that the fall of microorganisms from the upper to lower respiratory tract cannot be defended or excluded by mucociliary transportation disorder due to underlying disease and smoking, or deterioration of physical status other than overt or silent aspiration.

Age Factors↗

Prevalence of pathogens in diabetic foot infection in South Indian type 2 diabetic patients.

AIM: To determine the prevalence of pathogens in diabetic foot infections, in relation to parameters like Wagner's grading, duration of diabetes and healing time. MATERIAL AND METHODS: A group of 654 (M:F, 433:221) type 2 diabetic patients with foot ulcers were studied. Specimens like pus, wound exudate and tissue were processed for smear for Gram's staining, aerobic and anaerobic culture, and biochemical identifications. RESULTS: In 654 diabetic patients, 728 pathogens were isolated. Aerobic pathogens were isolated in 437 (66.8%) patients and anaerobic pathogens were isolated in 217 (33.2%). As Wagner's grading increased, the prevalence of anaerobic pathogens also increased. Neuropathy was common in diabetic patients infected with both aerobic and anaerobic pathogens. Ulcers infected with anaerobic pathogens showed a longer healing time than ulcers infected with aerobic pathogens. There was no significant difference in peripheral vascular disease (PVD) in patients selected for the study. Among aerobic pathogens, Enterobacteriaceae family (48%), Staphylococcus species (spp) (18.2%), Streptococcus spp (16.8%) and Pseudomonas spp (17%) were seen frequently. Among anaerobes Peptostreptococcus spp and Clostridium spp formed 69.4%. Gram-negative anaerobes like Bacteroides spp and Fusobacterium spp were present in 30.6%. Healing time was longer when strict aerobic pathogen Pseudomonas spp and strict anaerobic pathogens were present (136.1 +/- 28.6 and 136.4 +/- 34.7 days, respectively). CONCLUSIONS: Diabetic foot infection is polymicrobial in nature. The healing time of wound infected with anaerobic pathogens was higher than those infected with aerobic pathogens. Anaerobic pathogens increased with the Wagner's grading. Presence of neuropathy increased the risk of foot infection.

Bacteria, Aerobic↗

[Enteropathogens associated with acute diarrheal disease in children]

OBJECTIVE: Aiming a better understanding of the distribution of enteropathogens in Brazil, we have investigated the prevalence of enteropathogens in acute infantile diarrhea in children of low economic level from São José do Rio Preto, SP.METHODS: A prospective study with 196 children under age five, all with diarrhea, and 33 control subjects was conducted at Hospital de Base de São José do Rio Preto during 1995. This Hospital is associated to a Medicine School and offers free assistance to patients of low economic level who live in the area. RESULTS: Enteropathogens were identified in 48% of the cases and 27% of the controls. Shigella species were isolated in 21% of the cases, EPEC in 10.7%, Salmonella species in 5.6%. The other enteropathogens studied represented 12 % of the isolate pathogens from case children. In the controls 18.2% of atypical EPEC were isolated. Isolation of Shigella species increased with increasing age of cases and peaked in spring, whereas EPEC was common in early infancy and peaked in spring and winter.CONCLUSIONS: Among São José do Rio Preto children, Shigella species and diarrheiogenic E. coli strains (EPEC, ETEC, EAggEC, EIEC) were isolated throughout the year as a cause of diarrhea bringing children to a medical attention. Most of the pathogens were isolated in spring (mainly Shigella and Salmonella), so the temperature and humidity of the environment must be very important.

Journal Article↗

[Pathogenicity of fungi isolated from American ginseng seeds and bioassay of fungicides against the pathogenic fungi].

OBJECTIVE: Detect the pathogenicity of fungi isolated from American ginseng seeds and select new fungicides against the pathogenic seed-born fungi. METHOD: inoculate the isolates to American ginseng seedlings to detect the pathogenicity, and seeded the hyphae disk at the center of agar plate containing chemicals to do fungicide bioassay. RESULT: Fusarium sp. isolate 5 and Alternara sp. isolate 13 had pathogenicity to American ginseng seedling. Sportak had the strongest inhibition to the two isolates, Carbendazim had better inhibition and Celest had weak inhibition. CONCLUSION: Sportak and Celest could be used as efficacious candidate chemicals to treat or coat the American ginseng seeds, and substitute Carbendazim, which has been used for many years, to control the diseases occurring in seedling stage of American ginseng.

Alternaria↗

Microbial causes of neonatal conjunctivitis.

We assessed the microbial causes of neonatal conjunctivitis by comparing 55 infants with purulent conjunctivitis and 60 healthy control infants. A mean of greater than 5 leukocytes per 1000X microscopic field was seen in Gram-stained smears obtained from the more inflamed eye in 77% of 30 untreated patients but none of 57 controls. Pathogens isolated more often from untreated patients than from controls included Haemophilus spp. (17% vs 2%, P = 0.01), Staphylococcus aureus (17% vs 2%, P = 0.01), Chlamydia trachomatis (14% vs 0%, P = 0.01), enterococci (8% vs 0%, P = 0.05), and Streptococcus pneumoniae (11% vs 2%, P = 0.06). One or more of these pathogens were isolated from the conjunctivae in 58% of patients and 5% of controls (P less than 0.001). Bacterial morphology seen on smear correlated with the pathogens cultured. Isolation of Haemophilus spp. or S. pneumoniae was associated with dacryostenosis. We conclude that several microbial pathogens are implicated in neonatal conjunctivitis. These organisms have differing susceptibilities to antimicrobial agents, so culture and sensitivity testing are required as a guide to therapy.

Bacterial Infections↗

Pathogenicity of recent isolates of infectious bursal disease virus in specific-pathogen-free chickens: protection conferred by an intermediate vaccine strain.

The pathogenicity of recent isolates of infectious bursal disease virus and the protection conferred against them by a commercial vaccine strain of intermediate virulence were examined in specific-pathogen-free chickens. Based on clinical signs, mortality, and macroscopic lesions in susceptible chickens, the isolates designated as A-Delmarva and U-28 were distinct from a previously known serotype I virulent isolate (Edgar). Histopathological analysis of the bursa of Fabricius did not establish differences between the field isolates. Although the vaccine strain produced some degree of bursal damage in antibody-free chickens, it was significantly less severe than the damage caused by the field isolates. The active immune response induced by vaccination was cross-protective against the pathological effects produced by the different isolates used in this study.

Animals↗

Nonpathogenic isolates of Yersinia enterocolitica do not contain functional inv-homologous sequences.

Previous studies have demonstrated a correlation between the ability of isolates of Yersinia enterocolitica to cause disease and to invade tissue culture cells in vitro. Two genes, inv and ail, isolated from a pathogenic strain of Y. enterocolitica have each been shown to confer this invasive phenotype upon Escherichia coli. Eighty pathogenic, invasive isolates studied by Miller et al. (Infect. Immun. 57:121-131, 1989) contained sequences homologous to both of these genes. Thirty-five nonpathogenic, noninvasive isolates similarly studied had no ail homology but carried inv-homologous sequences. We investigated inv-homologous sequences from four nonpathogenic isolates. Recombinant clones of these inv-homologous sequences did not confer the invasive phenotype upon E. coli. No RNA transcripts capable of encoding a full-length Inv protein were detected in the four noninvasive Yersinia strains. When the inv gene from a pathogenic isolate was introduced into two of these strains, the resulting transformants invaded tissue culture cells in vitro. The inv gene was transcribed in a pathogenic Yersinia isolate grown at 30 degrees C but not at all in these cells grown at 37 degrees C. The production of RNA transcripts homologous to inv in transformants was not regulated by temperature to the same degree as was seen for pathogenic isolates. We conclude that the inv gene in nonpathogenic strains of Y. enterocolitica is nonfunctional. Y. enterocolitica isolates epidemiologically linked to disease contain both a functional inv gene and a functional ail gene. Environmental isolates not associated with disease have a nonfunctional inv gene and no ail gene.

Gene Expression Regulation, Bacterial↗

The value of a CT-guided fine needle aspirate in infants with lung abscess.

OBJECTIVE: To describe the range of pathogens isolated from a lung abscess in infants less than one year of age. To assess the role of direct culture from the abscess. METHODS: The two index cases were managed in 2002. An institution-based review was conducted of all infants up to one year of age diagnosed with a lung abscess between 1989 and 2002. Data sources were hospital's disease index and Neonatal Intensive Care Unit Audit database using ICD9 and ICD10 diagnostic codes for 'lung abscess'. RESULTS: Five infants, under the age of one year, were treated for a lung abscess. In the one case where the abscess was left-sided it was associated with a congenital cystic adenomatoid malformation of the lung. Pathogens were isolated following direct culture of the abscess in four cases. In three cases a single pathogen was isolated: pseudomonas aeruginosa, staphylococcus aureus and haemophilus influenzae. In one case a mixture of escherichia coli, streptococcus milleri and an anaerobe, propionibacteria, were cultured. Antibiotic therapy was directed at the identified pathogen(s) in all four cases. There was no mortality or recurrence. CONCLUSION: Predisposing factors for a lung abscess in infancy include prematurity, assisted ventilation, congenital lung anomaly and aspiration. Given the range of potential pathogens, direct culture by CT-guided fine needle aspiration is recommended to direct appropriate intravenous medical therapy provided the abscess is located peripherally.

Anti-Bacterial Agents↗

Viral and bacterial diseases of Atlantic cod Gadus morhua, their prophylaxis and treatment: a review.

This review summarises the state of knowledge of both viral and bacterial diseases of Atlantic cod Gadus morhua, and their diagnosis, prophylaxis and treatment. The most important losses have been at the larval and juvenile stages, and vibriosis has long been the most important bacterial disease in cod, with Listonella (Vibrio) anguillarum dominant among pathogenic isolates. Vaccination of cod against pathogens such as L. anguillarum and Aeromonas salmonicida clearly demonstrates that the cod immune system possesses an effective memory and appropriate mechanisms sufficient for protection, at least against some diseases. Well-known viruses such as the nodavirus that causes viral encephalopathy and retinopathy (VER), infectious pancreatic necrosis virus (IPNV) and viral haemorrhagic septicaemia virus (VHSV) have been isolated from Atlantic cod and can be a potential problem under intensive rearing conditions. No commercial vaccines against nodavirus are currently available, whereas vaccines against IPNV infections based upon inactivated virus as well as IPNV recombinant antigens are available. A number of investigations of the pharmacokinetic properties of antibacterial agents in cod and their efficacy in treating bacterial infections have been reviewed.

Animals↗

In vivo evaluation of the pathogenicity of field isolates of infectious bronchitis virus.

The pathogenicity of 13 field isolates of infectious bronchitis virus (IBV) isolated from Georgia broiler farms from 1989 to 1992 was evaluated using the IBV and Escherichia coli mixed-infection model. Based on the clinical signs, mortality, and lesions, the isolates were classified as high, intermediate, and low in pathogenicity. The in vivo classification was compared with the serotype classification results obtained by reverse transcriptase-polymerase chain reaction-restriction fragment length polymorphism analysis. The high-pathogenicity group was composed of five isolates representing three serotypes: Arkansas, Georgia variant (GAV), and Massachusetts. Isolates in the intermediate- and low-pathogenicity groups were all representatives of the Connecticut serotype, except for one isolate, which belonged to the Massachusetts serotype.

Animals↗

Cell membrane permeability and mitochondrial dysfunction-inducing activities in cell-free supernatants from Serpulina hyodysenteriae serotypes 1 and 2.

Membrane permeability (MP) and mitochondrial dysfunction-inducing (MDI) activities were detected in cell-free supernatants (CFS) of Serpulina hyodysenteriae, using either hemoglobin release from porcine red blood cells (RBC) or cytoplasmic lactate dehydrogenase release from porcine peripheral blood lymphocytes (PBL), and reduction of the 3-(4,5-dimethylthiazoyl-2-y1)-2,5-diphenyltetrazolium bromide dye by porcine PBL. The MP and MDI activities of CFS correlated with each other for serotype 1 and 2 isolates taken at different population densities; however, the kinetics of toxin production varied between each serotype. The loss of enteropathogenicity of two field isolates with nonpathogenic phenotypes and pathogenic isolates passaged up to 45 times in vitro was not attributable to a loss of either membrane permeability or mitochondrial dysfunction-inducing activity of cell-free supernatants. Results from this study suggested the potential for two separate toxins being involved in the pathogenesis of swine dysentery, with the MDI activity correlating with age susceptibility to clinical disease.

Age Factors↗

Yersinia spp. and numbers, with particular reference to Y. enterocolitica bio/serotypes, occurring on Irish meat and meat products, and the influence of alkali treatment on their isolation.

Samples (n = 340) of Irish meat and meat products were examined to determine the incidence and initial numbers of Yersinia spp. present. In cases where no typical Yersinia colonies were observed, samples were cold enriched up to 21 days at 4 degrees C and streaked out on CIN agar with and without post-enrichment alkali treatment. The highest isolation rates of Yersinia were found using post-enrichment alkali treatment and an incubation temperature of 37 degrees C. All Yersinia isolates were identified to species level and Y. enterocolitica isolates were biotyped and serotyped. Pathogenic isolates (biotypes 1B-5) were examined for the presence of a virulence plasmid using dye binding reactions and agglutination tests. The incidence of Yersinia spp. was high on raw (89%) and cooked meats (49%) obtained fromdelicatessen counters but lower in pre-packaged cooked meats (18.8%). Initial numbers of Yersinia were generally < 2.7 log10cfu/g on raw meats and < 0.7 log10cfu/g on cooked.

Food Microbiology↗

Genetics and genomics in infectious disease susceptibility.

The past decade has witnessed a rapid transition from the first positional cloning of an infectious disease susceptibility gene (Slc11a1, also called Nramp1) in the mouse to genome-wide scans in human multicase families and the identification of potential disease-causing genes by simple inspection of the public human genome databases. Pathogen genome projects have facilitated multilocus sequence typing of pathogen isolates and studies of ecological fitness and virulence patterns in disease-causing isolates. Comparative sequence analysis of pathogen strains and functional genomics studies are now underway, hopefully providing new insight into infectious disease susceptibility.

Cloning, Molecular↗

Routine subcultures shown to be unnecessary in radiometric detection of bacteremia using three media.

A study to determine the necessity of routine blind subcultures when using the radiometric BACTEC-460 instrument for detection of bacteremia was conducted. In the first phase, blind subcultures were performed on day 10 of incubation; in the second phase, blind subcultures were performed on day 4; in the final phase the subculture was performed on day one. During the study, 6,208 blood cultures from 2,455 patients were inoculated into three different culture media. There were 454 microbial isolates recovered, including 269 presumed pathogenic isolates. The single significant discrepancy between blind subculture and radiometric detection was a case of Staphylococcus epidermidis septicemia in which the organism was detected 48 hours earlier by blind subculture. There were no cultures in which significant bacterial isolates were detected by blind subcultures but not radiometrically. Thus, under the conditions of this study, routine blind subcultures are not necessary when using the BACTEC radiometric system for detection of bacteremia.

Adult↗

Role of Pantoea agglomerans in opportunistic bacterial seed and boll rot of cotton (Gossypium hirsutum) grown in the field.

AIMS: To investigate the aetiology of seed and boll rot of cotton grown in South Carolina (SC). METHODS AND RESULTS: Bacteria were isolated from diseased locules of cotton bolls collected in a field in SC, USA and tested for the ability to cause comparable disease symptoms in greenhouse grown cotton fruit. Spontaneously generated rifampicin-resistant (Rif(r)) mutants of the isolates were used in confirmatory pathogenicity tests. Resistance to the antibiotic was both stable and effective in differentiating between an inoculated Rif(r) strain, rifampicin-sensitive contaminants and/or endophytes. A series of inoculation methods was tested at various boll developmental stages and at different fruiting nodes on the plant. Field disease symptoms were reproduced by inoculating bolls at 2 weeks postanthesis with bacterial suspensions ranging from 10(3) to 10(6) CFU ml(-1). Pathogenic isolates were categorized as Pantoea agglomerans on the basis of phenotype testing, fatty acid profiling (similarity index = 0.94), and 16s ribosomal DNA sequence analysis (99% nucleotide identity). CONCLUSIONS: Pantoea agglomerans isolates from field-collected immature, diseased cotton caused comparable infection symptoms in greenhouse produced cotton fruit. SIGNIFICANCE AND IMPACT OF THE STUDY: In 1999, significant yield losses in SC cotton resulted from a previously unobserved seed and boll rot that has since been reported in other southeastern states. This study demonstrated a role of P. agglomerans in producing opportunistic bacterial seed and boll rot of cotton.

Anti-Bacterial Agents↗

Amoxycillin-clavulanic acid (Augmentin) versus metronidazole as prophylaxis in hysterectomy: a prospective, randomized clinical trial.

In order to determine the most effective regimen for the prevention of infection after elective hysterectomy, 300 patients were randomly assigned to receive three perioperative doses of either amoxycillin-clavulanic acid (1.2 g intravenous) or metronidazole (1 g suppository). Of the 280 patients who were assessable 138 were given amoxycillin-clavulanic acid and 142 received metronidazole; 268 underwent abdominal hysterectomy and 12 had vaginal hysterectomy. Patients in the amoxycillin-clavulanic acid group had significantly less infectious morbidity (13.8%) than those in the metronidazole group (33.1%). There were also statistically significant differences in favour of amoxycillin-clavulanic acid with respect to operative site infection, duration of hospital stay, need for postoperative antimicrobials, and surgery for operative site infection. But for one isolate of Bacteroides fragilis, all pathogens isolated from wound infections in the metronidazole group were aerobes. No anaerobes were isolated from patients in the amoxycillin-clavulanic acid group. The results suggest that prophylaxis for hysterectomy should consist of an agent, or combination of agents, with activity against both aerobic and anaerobic bacteria. Amoxycillin-clavulanic acid fulfils this criterion and appears to be effective and safe.

Adult↗

Enteric media: pre-poured or in-house?

AIMS: (1) To evaluate the performance of in-house and pre-poured commercially available enteric agar by challenge with a large number of positive clinical specimens. (2) To set the standard (critical independent evaluation) which new products should reach. (3) To publish this information, so that others can make informed decisions about enteric media. METHODS: Thirteen media of anonymous source were challenged with "known" positive stool samples. RESULTS: In-house desoxycholate citrate agar performed best for overall pathogen isolation rates, for shigella isolation rates, and for most pathogens available on primary culture. CONCLUSIONS: Desoxycholate citrate agar made by our own laboratory yielded the most pathogens and proved the most effective.

Agar↗