bluuebunny/crossref_metadata_2025 huggingface dataset:
Effect of Fermented Feed on the Microbial Population of the Gastrointestinal Tracts of Pigs
Authors: van Winsen René L. , Urlings Bert A. P. , Lipman Len J. A. , Snijders Jos M. A. , Keuzenkamp David , Verheijden Jos H. M. , van Knapen Frans
Published: July 2002

An in vivo experiment was performed with pigs to study the inhibitory effect of fermented feed on the bacterial population of the gastrointestinal tract. Results demonstrated a significant positive correlation between pH and lactobacilli in the stomach contents of pigs in dry feed as well as in the stomach contents of pigs fed fermented feed. Furthermore, a significant positive correlation between the pH and the numbers of bacteria in the family
Enterobacteriaceae
in the contents of the stomach of pigs fed dry feed was found. In the stomach contents of pigs fed fermented feed, a significant negative correlation was found between the concentration of the undissociated form of lactic acid and the numbers of
Enterobacteriaceae
. The numbers of
Enterobacteriaceae
in the contents of the stomach, ileum, cecum, colon, and rectum of pigs fed fermented feed were significantly lower compared with the contents of the stomach, ileum, caecum, colon, and rectum of pigs fed dry feed. The numbers of total lactobacilli were significantly higher in the stomach contents of pigs fed fermented feed and in the ileum contents of one pig group fed fermented feed compared with the contents of pigs fed dry feed. However, the influence of lactobacilli on numbers of
Enterobacteriaceae
could not be demonstrated. It was concluded that fermented feed influences the bacterial ecology of the gastrointestinal tract and reduces the levels of
Enterobacteriaceae
in the different parts of the gastrointestinal tract.
Fate of the Biological Control Agent
Pseudomonas aureofaciens
TX-1 after Application to Turfgrass
Authors: Sigler William V. , Nakatsu Cindy H. , Reicher Zachary J. , Turco Ronald F.
Published: July 2002

The fate and impact of
Pseudomonas aureofaciens
TX-1 following application as a biocontrol agent for fungi in turfgrass were studied. The organism was applied with a modified irrigation system by using a preparation containing 1 × 10
6
P. aureofaciens
TX-1 CFU ml
−1
about 100 times between May and August. We examined the impact of this repeated introduction of
P. aureofaciens
TX-1 (which is known to produce the antimicrobial compound phenazine-1-carboxylic acid) on the indigenous microbial community of the turfgrass system and on establishment of introduced bacteria in the soil system. A PCR primer-DNA hybridization probe combination was developed to accurately monitor the fate of
P. aureofaciens
TX-1 following application in irrigation water. To assess the impact of frequent
P. aureofaciens
TX-1 applications on the indigenous bacterial community, turfgrass canopy, thatch, and rhizosphere samples were obtained during the growing season from control and treated plots and subjected to DNA extraction procedures and denaturing gradient gel electrophoresis (DGGE). PCR amplification and hybridization of extracted DNA with the
P. aureofaciens
TX-1-specific primer-probe combination revealed that
P. aureofaciens
TX-1 not only became established in the rhizosphere and thatch but also was capable of overwintering. Separation of PCR-amplified partial 16S rRNA genes by DGGE showed that the repeated application of
P. aureofaciens
TX-1 in irrigation water resulted in transient displacement of a leaf surface bacterial community member. There was no obvious alteration of any dominant members of the thatch and rhizosphere microbial communities.
Bacillus
Spore Inactivation Methods Affect Detection Assays
Authors: Dang Jessica L. , Heroux Karen , Kearney John , Arasteh Ameneh , Gostomski Mark , Emanuel Peter A.
Published: July 2002

Detection of biological weapons is a primary concern in force protection, treaty verification, and safeguarding civilian populations against domestic terrorism. One great concern is the detection of
Bacillus anthracis
, the causative agent of anthrax. Assays for detection in the laboratory often employ inactivated preparations of spores or nonpathogenic simulants. This study uses several common biodetection platforms to detect
B. anthracis
spores that have been inactivated by two methods and compares those data to detection of spores that have not been inactivated. The data demonstrate that inactivation methods can affect the sensitivity of nucleic acid- and antibody-based assays for the detection of
B. anthracis
spores. These effects should be taken into consideration when comparing laboratory results to data collected and assayed during field deployment.
Characterization by 16S rRNA Sequence Analysis of Pseudomonads Causing Blotch Disease of Cultivated
Agaricus bisporus
Authors: Godfrey S. A. C. , Harrow S. A. , Marshall J. W. , Klena J. D.
Published: July 2002

Bacterial blotch of
Agaricus bisporus
has typically been identified as being caused by either
Pseudomonas tolaasii
(brown blotch) or
Pseudomonas gingeri
(ginger blotch). To address the relatedness of pseudomonads able to induce blotch, a pilot study was initiated in which pseudomonads were selectively isolated from mushroom farms throughout New Zealand. Thirty-three pseudomonad isolates were identified as being capable of causing different degrees of discoloration (separable into nine categories) of
A. bisporus
tissue in a bioassay. These isolates were also identified as unique using repetitive extragenic palindromic PCR and biochemical analysis. Relationships between these 33 blotch-causing organisms (BCO) and a further 22 selected pseudomonad species were inferred by phylogenetic analyses of near-full-length 16S rRNA gene nucleotide sequences. The 33 BCO isolates were observed to be distributed throughout the
Pseudomonas fluorescens
intrageneric cluster. These results show that in addition to known BCO (
P. tolaasii, P. gingeri
, and
Pseudomonas reactans
), a number of diverse pseudomonad species also have the ability to cause blotch diseases with various discolorations. Furthermore, observation of ginger blotch discoloration of
A. bisporus
being independently caused by many different pseudomonad species impacts on the homogeneity and classification of the previously described
P. gingeri
.
16S rRNA-Based Analysis of Microbiota from the Cecum of Broiler Chickens
Authors: Zhu Xiang Y. , Zhong Tanya , Pandya Yoga , Joerger Rolf D.
Published: July 2002

The microbiota of the intestinal tract of chickens plays an important role in inhibiting the establishment of intestinal pathogens. Earlier culturing and microscopic examinations indicated that only a fraction of the bacteria in the cecum of chickens could be grown in the laboratory. Therefore, a survey of cecal bacteria was done by retrieval of 16S rRNA gene sequences from DNA isolated from the cecal content and the cecal mucosa. The ribosomal gene sequences were amplified with universal primers and cloned or subjected to temporal temperature gradient gel electrophoresis (TTGE). Partial 16S rRNA gene sequences were determined from the clones and from the major bands in TTGE gels. A total of 1,656 partial 16S rRNA gene sequences were obtained and compared to sequences in the GenBank. The comparison indicated that 243 different sequences were present in the samples. Overall, sequences representing 50 phylogenetic groups or subgroups of bacteria were found, but approximately 89% of the sequences represented just four phylogenetic groups (
Clostridium leptum
,
Sporomusa
sp.,
Clostridium coccoides
, and enterics). Sequences of members of the
Bacteroides
group, the
Bifidobacterium infantis
subgroup, and of
Pseudomonas
sp. each accounted for less than 2% of the total. Sequences related to those from the
Escherichia
sp. subgroup and from
Lactobacillus
,
Pseudomonas
, and
Bifidobacterium
spp
.
were generally between 98 and 100% identical to sequences already deposited in the GenBank. Sequences most closely related to those of the other bacteria were generally 97% or less identical to those in the databases and therefore might be from currently unknown species. TTGE and random cloning indicated that certain phylogenetic subgroups were common to all birds analyzed, but sequence data from random cloning also provided evidence for qualitative and quantitative differences among the cecal microbiota of individual birds reared under very similar conditions.
Salt Stress Proteins Induced in
Listeria monocytogenes
Authors: Duché Ophélie , Trémoulet Frédéric , Glaser Philippe , Labadie Jean
Published: July 2002

The ability of
Listeria monocytogenes
to tolerate salt stress is of particular importance, as this pathogen is often exposed to such environments during both food processing and food preservation. In order to understand the survival mechanisms of
L. monocytogenes
, an initial approach using two-dimensional polyacrylamide gel electrophoresis was performed to analyze the pattern of protein synthesis in response to salt stress. Of 400 to 500 visible proteins, the synthesis of 40 proteins (
P
< 0.05) was repressed or induced at a higher rate during salt stress. Some of the proteins were identified on the basis of mass spectrometry or N-terminal sequence analysis and database searching. Twelve proteins showing high induction after salt stress were similar to general stress proteins (Ctc and DnaK), transporters (GbuA and mannose-specific phosphotransferase system enzyme IIAB), and general metabolism proteins (alanine dehydrogenase, CcpA, CysK, EF-Tu, Gap, GuaB, PdhA, and PdhD).
Effects of Growth Mode and Pyruvate Carboxylase on Succinic Acid Production by Metabolically Engineered Strains of
Escherichia coli
Authors: Vemuri G. N. , Eiteman M. A. , Altman E.
Published: July 2002

Escherichia coli
NZN111, which lacks activities for pyruvate-formate lyase and lactate dehydrogenase, and AFP111, a derivative which contains an additional mutation in
ptsG
(a gene encoding an enzyme of the glucose phophotransferase system), accumulate significant levels of succinic acid (succinate) under anaerobic conditions. Plasmid pTrc99A-
pyc
, which expresses the
Rhizobium etli
pyruvate carboxylase enzyme, was introduced into both strains. We compared growth, substrate consumption, product formation, and activities of seven key enzymes (acetate kinase, fumarate reductase, glucokinase, isocitrate dehydrogenase, isocitrate lyase, phosphoenolpyruvate carboxylase, and pyruvate carboxylase) from glucose for NZN111, NZN111/pTrc99A-
pyc
, AFP111, and AFP111/pTrc99A-
pyc
under both exclusively anaerobic and dual-phase conditions (an aerobic growth phase followed by an anaerobic production phase). The highest succinate mass yield was attained with AFP111/pTrc99A-
pyc
under dual-phase conditions with low pyruvate carboxylase activity. Dual-phase conditions led to significant isocitrate lyase activity in both NZN111 and AFP111, while under exclusively anaerobic conditions, an absence of isocitrate lyase activity resulted in significant pyruvate accumulation. Enzyme assays indicated that under dual-phase conditions, carbon flows not only through the reductive arm of the tricarboxylic acid cycle for succinate generation but also through the glyoxylate shunt and thus provides the cells with metabolic flexibility in the formation of succinate. Significant glucokinase activity in AFP111 compared to NZN111 similarly permits increased metabolic flexibility of AFP111. The differences between the strains and the benefit of pyruvate carboxylase under both exclusively anaerobic and dual-phase conditions are discussed in light of the cellular constraint for a redox balance.
Quantification in Soil and the Rhizosphere of the Nematophagous Fungus
Verticillium chlamydosporium
by Competitive PCR and Comparison with Selective Plating
Authors: Mauchline T. H. , Kerry B. R. , Hirsch P. R.
Published: July 2002

A competitive PCR (cPCR) assay was developed to quantify the nematophagous fungus
Verticillium chlamydosporium
in soil. A γ-irradiated soil was seeded with different numbers of chlamydospores from
V. chlamydosporium
isolate 10, and samples were obtained at time intervals of up to 8 weeks. Samples were analyzed by cPCR and by plating onto a semiselective medium. The results suggested that saprophytic
V. chlamydosporium
growth did occur in soil and that the two methods detected different phases of growth. The first stage of growth, DNA replication, was demonstrated by the rapid increase in cPCR estimates, and the presumed carrying capacity (PCC) of the soil was reached after only 1 week of incubation. The second stage, an increase in fungal propagules presumably due to cell division, sporulation, and hyphal fragmentation, was indicated by a less rapid increase in CFU, and 3 weeks was required to reach the PCC. Experiments with field soil revealed that saprophytic fungal growth was limited, presumably due to competition from the indigenous soil microflora, and that the PCR results were less variable than the equivalent plate count results. In addition, the limit of detection of
V. chlamydosporium
in field soil was lower than that in γ-irradiated soil, suggesting that there was a background population of the fungus in the field, although the level was below the limit of detection. Tomatoes were infected with the root knot nematode (RKN) or the potato cyst nematode (PCN) along with a PCN-derived isolate of the fungus (
V. chlamydosporium
isolate Jersey). Increases in fungal growth were observed in the rhizosphere of PCN-infested plants but not in the rhizosphere of RKN-infested plants after 14 weeks using cPCR. In this paper we describe for the first time PCR-based quantification of a fungal biological control agent for nematodes in soil and the rhizosphere, and we provide evidence for nematode host specificity that is highly relevant to the biological control efficacy of this fungus.
Use of the
Verrucomicrobia
-Specific Probe EUB338-III and Fluorescent In Situ Hybridization for Detection of “
Candidatus
Xiphinematobacter” Cells in Nematode Hosts
Authors: Vandekerckhove Tom T. M. , Coomans August , Cornelis Karen , Baert Philippe , Gillis Monique
Published: July 2002

Fluorescent in situ hybridization with a 16S rRNA probe specific for
Verrucomicrobia
was used to (i) confirm the division-level identity of and (ii) study the behavior of the obligate intracellular verrucomicrobium “
Candidatus
Xiphinematobacter” in its nematode hosts. Endosymbionts in the egg move to the pole where the gut primordium arises; hence, they populate the intestinal epithelia of juvenile worms. During the host's molt to adult female, the endosymbionts concentrate around the developing ovaries to occupy the ovarian wall. Some bacteria are enclosed in the ripening oocytes for vertical transmission.
Verrucomicrobia
in males stay outside the testes because the tiny spermatozoids are not suitable for transmission of cytoplasmic bacteria.
Identification of Nontuberculous Mycobacteria Existing in Tap Water by PCR-Restriction Fragment Length Polymorphism
Authors: Chang Chiao-tang , Wang Ling-yu , Liao Chen-yi , Huang Shiao-ping
Published: July 2002

This paper presents the finding of the possible cause of the high false-positive rate in acid-fast staining in histological examinations. Using acid-fast staining, culture, and PCR, acid-fast bacilli were detected in 83.7% of 49 hospital tap water samples and nontuberculous mycobacteria (NTM) were detected in 20.4% of the same 49 samples. The 10 NTM isolates were also identified to the species level using PCR-restriction fragment length polymorphism. Our findings indicate that NTM in hospital tap water are the possible cause of false positives in acid-fast staining and of nosocomial infection in immunocompromised patients.
Down Regulation of CD4 Expression following Isolation and Culture of Human Monocytes
Authors: Graziani-Bowering Gina M. , Filion Lionel G.
Published: July 2002

The down regulation of CD4 by cultured monocytes has been observed by our group and by other investigators. Flow cytometric experiments were done to examine which factors might influence this phenomenon. The addition of lipopolysaccharide, granulocyte-macrophage colony-stimulating factor, macrophage colony-stimulating factor, or interleukin-10 to monocyte cultures failed to inhibit the decrease in monocyte CD4 expression routinely observed following overnight culture. The down regulation was an adherence-independent phenomenon and was not influenced by the type of anticoagulant into which the peripheral blood was collected or by the presence or absence of lymphocytes within the cultures. The avoidance of the use of Ficoll-Paque to isolate peripheral blood mononuclear cells did not prevent monocyte CD4 down regulation. Finally, by tagging monocyte CD4 with an anti-CD4 phycoerythrin-conjugated monoclonal antibody prior to culture, we were able to determine that the down regulation observed was the result of the internalization of the molecule. At this time, we conclude that the observed down regulation of monocyte CD4 is probably due to the differentiation of blood monocytes into tissue culture-derived macrophages rather than to some artifact of the isolation procedure.
Three-Color Flow Cytometry Detection of Intracellular Cytokines in Peripheral Blood Mononuclear Cells: Comparative Analysis of Phorbol Myristate Acetate-Ionomycin and Phytohemagglutinin Stimulation
Authors: Baran Jarołsaw , Kowalczyk Danuta , Oz˙óg Mariola , Zembala Marek
Published: July 2002

The assessment of intracellular cytokines at the single-cell level by flow cytometry has recently become a potent tool in many areas of cell biology and in defining the role of cytokines in various human diseases. Three-color flow cytometry for detection of intracellular cytokines combined with simultaneous determination of lymphocytes (CD3
+
and CD4
+
) or monocytes (CD33
+
and CD14
+
) was used for comparison of phytohemagglutinin (PHA)-and phorbol myristate acetate (PMA)-ionomycin-induced production of intracellular cytokines in peripheral blood mononuclear cells (PBMCs) of healthy donors. We found that the number of PBMCs stained for tumor necrosis factor alpha and gamma interferon after 6 h of activation was higher when PMA-ionomycin was used for stimulation, while the frequencies of cells positive for interleukin 4 (IL-4) were similar for both stimulators. However, PMA-ionomycin stimulation caused prominent alterations of cell morphology and membrane expression of CD4 and CD14. In contrast, PHA did not cause downregulation of surface markers and resulted in less pronounced alterations in both forward and side scatter signals during flow cytometry analysis. Moreover, during 48 h of culture PHA stimulated tumor necrosis factor beta and IL-10 production, which was not observed when PMA-ionomycin was used. We conclude that the use of PHA for cell activation may limit in vitro artifacts and allow more precise analysis of intracellular cytokine production in various disease states.
Rhesus Monkey (
Macaca mulatta
) Mucosal Antimicrobial Peptides Are Close Homologues of Human Molecules
Authors: Bals Robert , Lang Christiane , Weiner Daniel J. , Vogelmeier Claus , Welsch Ulrich , Wilson James M.
Published: July 2002

One component of host defense at mucosal surfaces appears to be epithelium-derived antimicrobial peptides. Molecules of the defensin and cathelicidin families have been studied in several species, including human and mouse. We describe in this report the identification and characterization of rhesus monkey homologues of human mucosal antimicrobial peptides. Using reverse transcriptase PCR methodology, we cloned the cDNAs of rhesus monkey β-defensin 1 and 2 (rhBD-1 and rhBD-2) and rhesus monkey LL-37/CAP-18 (rhLL-37/rhCAP-18). The predicted amino acid sequences showed a high degree of homology to the human molecules. The expression of the monkey antimicrobial peptides was analyzed using immunohistochemistry with three polyclonal antibodies to the human molecules. As in humans, rhesus monkey antimicrobial peptides are expressed in epithelia of various organs. The present study demonstrates that β-defensins and cathelicidins of rhesus monkeys are close homologues to the human molecules and indicate that nonhuman primates represent valid model organisms to study innate immune functions.
Biological Properties of Lipid A Isolated fromFlavobacterium meningosepticum
Authors: Tanamoto Ken-Ichi , Kato Hitomi , Haishima Yuji , Azumi Satoko
Published: July 2002

The biological properties of the lipid A fromFlavobacterium meningosepticum, which we recently isolated and whose complete chemical structure has been determined (H. Kato, T. Iida, Y. Haishima, A. Tanaka, and K. Tanamoto. J. Bacteriol. 180:3891–3899, 1998), were studied. The lipid A exhibited generally moderate activity compared toSalmonella entericasubsp.entericaserovar abortus equi lipopolysaccharide (LPS) used as a control in the assay systems tested; lethal toxicity in galactosamine-sensitized mice, mitogenicity in mouse spleen cells, induction of tumor necrosis factor alpha (TNF-α) release from mouse peritoneal macrophages and J774-1 mouse macrophage-like and human THP-1 line cells, nitric oxide induction activity from J774-1 cells, andLimulusgelation activity. The moderate activity of theF. meningosepticumlipid A may be explained by its unique fatty acid composition and the lack of a phosphate group in position 4′. It is noteworthy that the lipid A apparently induced TNF-α release from peritoneal macrophages in LPS-unresponsive C3H/HeJ mice and that the activation was suppressed by the LPS-specific antagonist, succinylated lipid A precursor. Significant splenocyte mitogenicity in C3H/HeJ mice was also observed with the lipid A. Taken together with the previous results concerningPorphyromonas gingivalislipid A, which has a high level of structural similarity to the lipid A ofF. meningosepticum, and the induction of TNF-α release in macrophages from C3H/HeJ mice, the lipid A ofF. meningosepticum, which has novel fatty acids, may possibly play an role for the activation of C3H/HeJ macrophages.
Persistence of
Lactobacillus fermentum
RC-14 and
Lactobacillus rhamnosus
GR-1 but Not
L. rhamnosus
GG in the Human Vagina as Demonstrated by Randomly Amplified Polymorphic DNA
Authors: Gardiner Gillian E. , Heinemann Christine , Bruce Andrew W. , Beuerman Dee , Reid Gregor
Published: July 2002

Lactobacillus rhamnosus
GR-1 and
L. fermentum
RC-14 are well-characterized probiotic strains with efficacy in the prevention and treatment of urogenital infections in women. The aim of the present study was to apply a molecular biology-based methodology for the detection of these strains and
L. rhamnosus
GG (a commercially available intestinal probiotic) in the human vagina in order to assess probiotic persistence at this site. Ten healthy women inserted vaginally a capsule containing either a combination of strains GR-1 and RC-14 or the GG strain for 3 consecutive nights. Vaginal swabs taken before and at various time points after probiotic insertion were analyzed, and the
Lactobacillus
flora was assessed by randomly amplified polymorphic DNA (RAPD) analysis. This method generated discrete DNA fingerprints for GR-1, RC-14, and GG and enabled successful detection of these strains in the vagina. Strain GR-1 and/or strain RC-14 was found to persist in the vaginal tract for up to 19 days after vaginal instillation, while
L. rhamnosus
GG was detectable for up to 5 days postadministration. In conclusion, the fates of probiotic
L. rhamnosus
and
L
.
fermentum
strains were successfully monitored in the human vagina by RAPD analysis. This technique provides molecular biology-based evidence that RC-14 and GR-1, strains selected as urogenital probiotics, persist in the human vagina and may be more suited to vaginal colonization than
L. rhamnosus
GG. This highlights the importance of proper selection of strains for urogenital probiotic applications.
Kinetics of Antibody Responses in
Rickettsia africae
and
Rickettsia conorii
Infections
Authors: Fournier Pierre-Edouard , Jensenius Mogens , Laferl Herman , Vene Sirka , Raoult Didier
Published: July 2002

African tick-bite fever, caused by
Rickettsia africae
, is the most common tick-borne rickettsiosis in sub-Saharan Africa. Mediterranean spotted fever due to
Rickettsia conorii
also occurs in the region but is more prevalent in Mediterranean countries. Using microimmunofluorescence, we compared the development of immunoglobulin G (IgG) and IgM titers in 48 patients with African tick-bite fever and 48 patients with Mediterranean spotted fever. Doxycycline treatment within 7 days from the onset of disease significantly prevented the development of antibodies to
R. africae
. In patients with African tick-bite fever, the median times to seroconversion with IgG and IgM were 28 and 25 days, respectively, after the onset of symptoms. These were significantly longer by a median of 6 days for IgG and 9 days for IgM than the times for seroconversion in patients with Mediterranean spotted fever (
P
< 10
−2
). We recommend that sera collected 4 weeks after the onset of signs of patients with suspected African tick-bite fever should be used for the definitive serological diagnosis of
R. africae
infections.
Hepatitis A: Old and New
Authors: Cuthbert Jennifer A.
Published: July 2002

The hepatitis A virus (HAV), a picornavirus, is a common cause of hepatitis worldwide. Spread of infection is generally person to person or by oral intake after fecal contamination of skin or mucous membranes; less commonly, there is fecal contamination of food or water. Hepatitis A is endemic in developing countries, and most residents are exposed in childhood. In contrast, the adult population in developed countries demonstrates falling rates of exposure with improvements in hygiene and sanitation. The export of food that cannot be sterilized, from countries of high endemicity to areas with low rates of infection, is a potentially important source of infection. After ingestion and uptake from the gastrointestinal tract, the virus replicates in the liver and is excreted into the bile. Cellular immune responses to the virus lead to destruction of infected hepatocytes with consequent development of symptoms and signs of disease. Humoral immune responses are the basis for diagnostic serologic assays. Acute HAV infection is clinically indistinguishable from other causes of acute viral hepatitis. In young children the disease is often asymptomatic, whereas in older children and adults there may be a range of clinical manifestations from mild, anicteric infection to fulminant hepatic failure. Clinical variants include prolonged, relapsing, and cholestatic forms. Management of the acute illness is supportive, and complete recovery without sequelae is the usual outcome. Research efforts during World War II led to the development of passive immunoprophylaxis. Pooled immune serum globulin is efficacious in the prevention and attenuation of disease in exposed individuals. More recently, active immunoprophylaxis by vaccination has been accomplished. Future eradication of this disease can now be contemplated.
Nosocomial Spread of Viral Disease
Authors: Aitken Celia , Jeffries Donald J.
Published: July 2002

Viruses are important causes of nosocomial infection, but the fact that hospital outbreaks often result from introduction(s) from community-based epidemics, together with the need to initiate specific laboratory testing, means that there are usually insufficient data to allow the monitoring of trends in incidences. The most important defenses against nosocomial transmission of viruses are detailed and continuing education of staff and strict adherence to infection control policies. Protocols must be available to assist in the management of patients with suspected or confirmed viral infection in the health care setting. In this review, we present details on general measures to prevent the spread of viral infection in hospitals and other health care environments. These include principles of accommodation of infected patients and approaches to good hygiene and patient management. They provide detail on individual viral diseases accompanied in each case with specific information on control of the infection and, where appropriate, details of preventive and therapeutic measures. The important areas of nosocomial infection due to blood-borne viruses have been extensively reviewed previously and are summarized here briefly, with citation of selected review articles. Human prion diseases, which present management problems very different from those of viral infection, are not included.
Vaccination of Cattle withMycobacterium bovisCulture Filtrate Proteins and Interleukin-2 for Protection against Bovine Tuberculosis
Authors: Wedlock D. Neil , Vesosky Bridget , Skinner Margot A. , de Lisle Geoffrey W. , Orme Ian M. , Buddle Bryce M.
Published: July 2002

In this study vaccines prepared from culture filtrate proteins (CFP) ofMycobacterium bovisand interleukin-2 (IL-2) were tested in cattle for their capacity to stimulate immune responses and to protect against an intratracheal challenge with virulentM. bovis. Nine groups of cattle were vaccinated with combinations of different doses of CFP and bovine IL-2 mixed with a monophosphoryl lipid A (MPL) adjuvant. An additional group was vaccinated withM. bovisBCG. Immune responses in CFP–IL-2-vaccinated animals differed from those seen in BCG-vaccinated animals by inducing high antigen-specific antibody responses and low levels of gamma interferon and IL-2 released from purified protein derivative-stimulated whole-blood cultures. In a concurrent experiment, additional animals were added to the high-dose CFP–IL-2, MPL control, and BCG groups and these expanded groups of animals were challenged intratracheally with virulentM. bovis. Although the lung lesion scores were significantly lower for both the CFP–IL-2-and BCG-vaccinated groups compared to the MPL control group, the overall level of protection was greatest for the BCG-vaccinated animals. There were more animals with extrathoracic spread of disease in the CFP–IL-2 group than in the other groups. While vaccination of cattle withM. bovisCFP gave an encouraging reduction in tuberculous lesions and did not induce a delayed-type hypersensitivity response to PPD, future CFP vaccines must prevent any extrathoracic spread of disease.
Analysis of Urease Expression in
Actinomyces naeslundii
WVU45
Authors: Morou-Bermudez Evangelia , Burne Robert A.
Published: July 2002

The hydrolysis of urea by ureases of oral bacteria in dental plaque can cause a considerable increase in plaque pH, which can inhibit the development of dental caries. There is also indirect evidence that urea metabolism may promote the formation of calculus and that ammonia release from urea could exacerbate periodontal diseases.
Actinomyces naeslundii
, an early colonizer of the oral cavity and a numerically significant plaque constituent, demonstrates comparatively low levels of urease activity on isolation, so this organism has not been considered a major contributor to total oral urease activity. In this study it was observed that urease activity and urease-specific mRNA levels in
A. naeslundii
WVU45 can increase up to 50-fold during growth under nitrogen-limiting conditions. Using primer extension analysis, a putative, proximal, nitrogen-regulated promoter of the
A. naeslundii
urease gene cluster was identified. The functionality and nitrogen responsiveness of this promoter were confirmed using reporter gene fusions and 5′ deletion analysis. The data indicated that regulation of urease expression by nitrogen availability in
A. naeslundii
may require a positive transcriptional activator. Plaque bacteria may experience nitrogen limitation when carbohydrates are present in excess. Therefore, based on the results of this study and in contrast to previous beliefs, strains of
A. naeslundii
may have the potential to be significant contributors to total plaque ureolysis, particularly during periods when there is an increased risk for caries development.
Identification of a
Porphyromonas gingivalis
Receptor for the
Streptococcus gordonii
SspB Protein
Authors: Chung Whasun O. , Demuth Donald R. , Lamont Richard J.
Published: July 2002

Colonization of the plaque biofilm by the oral pathogen
Porphyromonas gingivalis
is favored by the presence of antecedent organisms such as
Streptococcus gordonii
. Coadhesion between
P. gingivalis
and
S. gordonii
can be mediated by the SspB protein of
S. gordonii
; however, the
P. gingivalis
cognate receptor for this protein has not been identified. In this study, we identified a surface protein of
P. gingivalis
that interacts with the SspB protein. Coprecipitation between
P. gingivalis
outer membrane proteins and purified SspB protein demonstrated that a 100-kDa
P. gingivalis
protein bound to SspB. The 100-kDa protein also bound to an engineered strain of
Enterococcus faecalis
that expresses the SspB protein on the cell surface. Monospecific polyclonal antibodies to the 100-kDa protein inhibited the binding between
P. gingivalis
and
S. gordonii
in a dose-dependent manner up to 86%. Amino acid sequencing of the 100-kDa protein showed homology to a protein previously identified as the
P. gingivalis
minor fimbria. The minor fimbrial protein may exist as a complex with a hemagglutinin-like protein since the genes encoding these proteins are adjacent on the chromosome and are cotranscribed. Thus, the
P. gingivalis
receptor for
S. gordonii
SspB is a 100-kDa protein that structurally may be a minor fimbria-protein complex and functionally effectuates coadhesion.
Neutralizing Antibodies to Adenylate Cyclase Toxin Promote Phagocytosis ofBordetella pertussisby Human Neutrophils
Authors: Weingart Christine L. , Mobberley-Schuman Paula S. , Hewlett Erik L. , Gray Mary C. , Weiss Alison A.
Published: July 2002

A previous study showed that opsonization with human immune serum could either promote or antagonize phagocytosis ofBordetella pertussisby human neutrophils depending on whether the bacteria expressed adenylate cyclase toxin. Opsonization of the wild-type strain inhibited phagocytosis relative to unopsonized controls. In contrast, mutants lacking adenylate cyclase toxin were efficiently phagocytosed when opsonized with human immune serum. In this study, we examined opsonization in the presence or absence of monoclonal antibodies to adenylate cyclase toxin. Addition of neutralizing monoclonal antibodies to adenylate cyclase toxin converted a serum that previously inhibited both attachment and phagocytosis of the wild-type strain to one that increased both attachment and phagocytosis compared to the no-serum control. Monoclonal antibodies that recognize the adenylate cyclase toxin but fail to neutralize activity were without effect. These results suggest that adenylate cyclase toxin inhibits both Fc receptor-mediated attachment and phagocytosis ofB. pertussisby neutrophils.
Expression of Chemokine Genes in Murine Macrophages Infected withOrientia tsutsugamushi
Authors: Cho Nam-Hyuk , Seong Seung-Yong , Huh Myung-Suk , Han Tae-Hee , Koh Young-Sang , Choi Myung-Sik , Kim Ik-Sang
Published: July 2002

Scrub typhus, caused byOrientia tsutsugamushiinfection, is characterized by local as well as systemic inflammatory manifestations. Inflammation is initiated byO. tsutsugamushi-infected macrophages and endothelial cells in the dermis. We investigated the regulation of chemokine induction in macrophage cell line J774A.1 in response toO. tsutsugamushiinfection. The mRNAs for macrophage inflammatory proteins 1α/β (MIP-1α/β), MIP-2, and macrophage chemoattractant protein 1 were induced within 30 min, and their levels showed a transitory peak for 3 to 12 h. However, the lymphotactin, eotaxin, gamma interferon-inducible protein 10, and T-cell activation gene 3 mRNAs were not detected by RNase protection assays. Heat-killedO. tsutsugamushiinduced a similar extent of chemokine responses. Induction of the chemokine genes was not blocked by the eukaryotic protein synthesis inhibitor cycloheximide, suggesting that de novo synthesis of host cell protein is not required for these transcriptional responses. The induction of chemokine mRNAs byO. tsutsugamushiwas blocked by the inhibitors of NF-κB activation. Furthermore,O. tsutsugamushiinduced the nuclear translocation and activation of NF-κB. These results demonstrate that heat-stable molecules ofO. tsutsugamushiinduce a subset of chemokine genes and that induction involves activation of the transcription factor NF-κB.
Use of Lambda Phage S and R Gene Products in an Inducible Lysis System for
Vibrio cholerae
- and
Salmonella enterica
Serovar Typhimurium-Based DNA Vaccine Delivery Systems
Authors: Jain Vivek , Mekalanos John J.
Published: July 2002

Novel methods for adapting DNA vaccine technology to the prevention of mucosal diseases are greatly needed. Here we show that regulated expression of phage lambda lysis genes S and R causes dramatic lysis of both
Vibrio cholerae
and
Salmonella enterica
serovar Typhimurium cells with concomitant release of plasmid DNA into the surrounding media. We also used single and double DNase mutant strains to show that secreted
V. cholerae
DNases can adversely affect the integrity of DNA molecules released upon lysis. These results suggest that incorporation of lambda SR-mediated lysis constructs and DNA stabilizing mutations into candidate live attenuated bacterial vaccines offers a promising approach for the development of effective mucosal DNA delivery vectors for humans.
Genetic Relationships between Clinical Isolates of
Streptococcus pneumoniae
,
Streptococcus oralis
, and
Streptococcus mitis
: Characterization of “Atypical” Pneumococci and Organisms Allied to
S. mitis
Harboring
S. pneumoniae
Virulence Factor-Encoding Genes
Authors: Whatmore Adrian M. , Efstratiou Androulla , Pickerill A. Paul , Broughton Karen , Woodard Geoffrey , Sturgeon Daniel , George Robert , Dowson Christopher G.
Published: July 2002

The oral streptococcal group (mitis phylogenetic group) currently consists of nine recognized species, although the group has been traditionally difficult to classify, with frequent changes in nomenclature over the years. The pneumococcus (
Streptococcus pneumoniae
), an important human pathogen, is traditionally distinguished from the most closely related oral streptococcal species
Streptococcus mitis
and
Streptococcus oralis
on the basis of three differentiating characteristics: optochin susceptibility, bile solubility, and agglutination with antipneumococcal polysaccharide capsule antibodies. However, there are many reports in the literature of pneumococci lacking one or more of these defining characteristics. Sometimes called “atypical” pneumococci, these isolates can be the source of considerable confusion in the clinical laboratory. Little is known to date about the genetic relationships of such organisms with classical
S. pneumoniae
isolates. Here we describe these relationships based on sequence analysis of housekeeping genes in comparison with previously characterized isolates of
S. pneumoniae
,
S. mitis
, and
S. oralis
. While most pneumococci were found to represent a closely related group these studies identified a subgroup of atypical pneumococcal isolates (bile insoluble and/or “acapsular”) distinct from, though most closely related to, the “typical” pneumococcal isolates. However, a large proportion of isolates, found to be atypical on the basis of capsule reaction alone, did group with typical pneumococci, suggesting that they have either lost capsule production or represent as-yet-unrecognized capsular types. In contrast to typical
S. pneumoniae
, isolates phenotypically identified as
S. mitis
and
S. oralis
, which included isolates previously characterized in taxonomic studies, were genetically diverse. While most of the
S. oralis
isolates did fall into a well-separated group,
S. mitis
isolates did not cluster into a well-separated group. During the course of these studies we also identified a number of potentially important pathogenic isolates, which were frequently associated with respiratory disease, that phenotypically and genetically are most closely related to
S. mitis
but which harbor genes encoding the virulence determinants pneumolysin and autolysin classically associated with
S. pneumoniae
.
Colonization of the Respiratory Tract by a Virulent Strain of Avian
Escherichia coli
Requires Carriage of a Conjugative Plasmid
Authors: Ginns C. A. , Benham M. L. , Adams L. M. , Whithear K. G. , Bettelheim K. A. , Crabb B. S. , Browning G. F.
Published: July 2002

The E3 strain of
E. coli
was isolated in an outbreak of respiratory disease in broiler chickens, and experimental aerosol exposure of chickens to this strain induced disease similar to that seen in the field. In order to establish whether the virulent phenotype of this strain was associated with carriage of particular plasmids, four plasmid-cured derivatives, each lacking two or more of the plasmids carried by the wild-type strain, were assessed for virulence. Virulence was found to be associated with one large plasmid, pVM01. Plasmid pVM01 was marked by introduction of the transposon Tn
phoA
, carrying kanamycin resistance, and was then cloned by transformation of
E. coli
strain DH5α. The cloned plasmid was then reintroduced by conjugation into an avirulent plasmid-cured derivative of strain E3 which lacked pVM01. The conjugant was shown to be as virulent as the wild-type strain E3, establishing that this plasmid is required for virulence following aerosol exposure. This virulence plasmid conferred expression of a hydroxamate siderophore, but not colicins, on both strain E3 and strain DH5α. Carriage of this plasmid was required for strain E3 to colonize the respiratory tracts of chickens but was not necessary for colonization of the gastrointestinal tract. However, the virulence plasmid did not confer virulence, or the capacity to colonize the respiratory tract, on strain DH5α. Thus, these studies have established that infection of chickens with
E. coli
strain E3 by the respiratory route is dependent on carriage of a conjugative virulence plasmid, which confers the capacity to colonize specifically the respiratory tract and which also carries genes for expression of a hydroxymate siderophore. These findings will facilitate identification of the specific genes required for virulence in these pathogens.
Roles of the Surface Layer Proteins of
Campylobacter fetus
subsp.
fetus
in Ovine Abortion
Authors: Grogono-Thomas R. , Dworkin J. , Blaser M. J. , Newell D. G.
Published: July 2002

The role of the surface (S)-layer proteins of
Campylobacter fetus
subsp.
fetus
has been investigated using an ovine model of abortion. Wild-type strain 23D induced abortion in up to 90% of pregnant ewes challenged subcutaneously. Isolates recovered from both dams and fetuses expressed S-layer proteins with variable molecular masses. The spontaneous S-layer-negative variant, strain 23B, neither colonized nor caused abortions in pregnant ewes. A series of isogenic
sapA
and
recA
mutants, derived from 23D, also were investigated in this model. A mutant (501 [
sapA recA
+
]) caused abortion in one of five challenged animals and was recovered from the placenta of a second animal. Another mutant (502 [
sapA recA
]) with no S-layer protein expression caused no colonization or abortions in challenged animals but caused abortion when administered intraplacentally. Mutants 600(2) and 600(4), both
recA
, had fixed expression of 97- and 127-kDa S-layer proteins, respectively. Two of the six animals challenged with mutant 600(4) were colonized, but there were no abortions. As expected, all five strains recovered expressed a 127-kDa S-layer protein. In contrast, mutant 600(2) was recovered from the placentas of all five challenged animals and caused abortion in two. Unexpectedly, one of the 16 isolates expressed a 127-kDa rather than a 97-kDa S-layer protein. Thus, these studies indicate that S-layer proteins appear essential for colonization and/or translocation to the placenta but are not required to mediate fetal injury and that S-layer variation may occur in a
recA
strain.
Candida albicansandCandida kruseiDifferentially Induce Human Blood Mononuclear Cell Interleukin-12 and Gamma Interferon Production
Authors: Xiong Jingbo , Kang Kefei , Liu Liming , Yoshida Yuichi , Cooper Kevin D. , Ghannoum Mahmoud A.
Published: July 2002

Protection againstCandidainfection involves both innate and acquired immune responses, and cytokines produced by monocytes during the innate response may modify the acquired immune response by T cells. We hypothesized thatCandidaspecies which differ in pathogenicity can differentially induce production of immunoregulatory cytokines by human monocytes, which in turn modify T cells for immune responses toCandida. To test this hypothesis, we examined the effects ofCandida albicansandCandida kruseion immunoregulatory cytokine production by human monocytes and gamma interferon (IFN-γ) production by peripheral blood mononuclear cells (PBMC). Purified monocytes were incubated with live or heat-killed strains ofC. albicansandC. kruseiat the optimalCandida/monocyte ratio of 0.5. Cytokines in the supernatants were measured by enzyme-linked immunosorbent assay. Our data demonstrated that liveC. albicansandC. kruseisignificantly induced interleukin-10 (IL-10), monocyte chemotactic factor 1, IL-1β, and tumor necrosis factor alpha production by monocytes relative to unstimulated monocytes. In contrast, unlikeC. krusei, pathogenic live strains ofC. albicansinduced no or only a minimal level of IL-12. The expression of IL-12 p40 mRNA levels by reverse transcription-PCR corroborated the IL-12 protein (p70) findings. In human PBMC, human blood monocytes were the major source of both IL-10 and IL-12 production in response toC. albicansandC. krusei. Upon activation of T cells in the presence ofCandida-modified monocytes and antigen-presenting cells, IL-12 production by PBMC treated withCandidaorganisms correlated strongly with the level of IFN-γ production by T cells. These results indicate that the virulence ofC. albicansmay be related to its ability to induce the monocytic type II cytokine IL-10, with a selective inhibition of IL-12 production, which may be responsible for the observed lack of T-cell IFN-γ and may restrain an effective type I immune response toCandida.