A global perspective on bacterial diversity in the terrestrial deep subsurface.

While recent efforts to catalogue Earth's microbial diversity have focused upon surface and marine habitats, 12-20 % of Earth's biomass is suggested to exist in the terrestrial deep subsurface, compared to ~1.8 % in the deep subseafloor. Metagenomic studies of the terrestrial deep subsurface have yielded a trove of divergent and functionally important microbiomes from a range of localities. However, a wider perspective of microbial diversity and its relationship to environmental conditions within the terrestrial deep subsurface is still required. Our meta-analysis reveals that terrestrial deep subsurface microbiota are dominated by Betaproteobacteria, Gammaproteobacteria and Firmicutes, probably as a function of the diverse metabolic strategies of these taxa. Evidence was also found for a common small consortium of prevalent Betaproteobacteria and Gammaproteobacteria operational taxonomic units across the localities. This implies a core terrestrial deep subsurface community, irrespective of aquifer lithology, depth and other variables, that may play an important role in colonizing and sustaining microbial habitats in the deep terrestrial subsurface. An in silico contamination-aware approach to analysing this dataset underscores the importance of downstream methods for assuring that robust conclusions can be reached from deep subsurface-derived sequencing data. Understanding the global panorama of microbial diversity and ecological dynamics in the deep terrestrial subsurface provides a first step towards understanding the role of microbes in global subsurface element and nutrient cycling.

Deletion polymorphism in the angiotensin-converting enzyme gene as a thrombophilic risk factor after hip arthroplasty.

Despite thromboprophylaxis, deep vein thrombosis is a common complication of major orthopedic surgery. Predisposing genetic risk factors are unknown. In this case-control study, we investigated the association of the insertion (I)/deletion (D) angiotensin converting enzyme (ACE) gene polymorphism, Factor V Leiden (R506Q) mutation, and 5,10 methylenetetrahydrofolate reductase (MTHFR) gene polymorphism with post-operative venous thrombosis in 85 patients who underwent elective total hip arthroplasty. The odds of a thrombotic event following hip surgery among subjects with the DD genotype of the ACE gene was increased more than 10-fold compared to subjects with the II genotype (odds ratio 11.7 [95% confidence interval 2.3-84.5]); it was increased 5-fold in subjects with the ID genotype compared to the II genotype (odds ratio 5.0 [95% confidence interval 1.1-34.9]). Mean plasma ACE level in control subjects not on ACE inhibitors at the time of study (n=43) was lowest in persons homozygous for the I allele (18.9+/-7.95 U/l), intermediate in patients with the ID genotype (31.6+/-10.8 U/l) and highest in subjects homozygous for the D allele (44.0+/-7.14 U/l). Mean plasma ACE level among cases was higher (33.0 U/l, n=25) than among controls (29.4 U/l, n=43) but this difference was not statistically significant. Neither the Factor V Leiden mutation nor MTHFR gene polymorphism increased the risk of thrombosis following hip replacement. These results demonstrate that the I/D ACE gene polymorphism is a potent risk factor for thrombosis in subjects undergoing total hip arthroplasty.

Comparison of enrichment methods for recovery and chick infectivity of chlorine-injured Salmonella enteritidis.

In recent years, several preenrichment media have been shown to be effective for use in the recovery of sublethally injured Salmonella organisms. Selective enrichment without preenrichment has resulted in a lower recovery of organisms, particularly with regard to injured or stressed salmonellae. The present experiments compared the ability of nonselective preenrichment followed by selective enrichment or direct selective enrichment alone to recover chlorine-injured Salmonella organisms. Additionally, the Salmonella detection limits of the two enrichment methods were compared with minimal infectious dose in neonatal chicks. In three experiments, Salmonella enteritidis cells were exposed to chlorine for specific times and subsequently cultured by using preenrichment followed by selective enrichment or selective enrichment alone. Simultaneously, neonatal chicks were orally challenged with S. enteritidis cells from each exposure time to chlorine. The results indicated a marginal, but significantly (P < 0.05) higher level of recovery of sublethally injured salmonellae by using nonselective preenrichment followed by selective enrichment, as compared to selective enrichment alone. Interestingly, both culture methods were capable of detecting injured S. enteritidis cells at levels incapable of infecting neonatal chicks.

Photodecomposition of 1,2,3,4- and 2,3,7,8-tetrachlorodibenzo-p-dioxin (TCDD) in water-alcohol media on a solid support.

We used a hydrophobic solid support, octadecylsilylated silica gel (C18), packed in a quartz column as a reaction medium for the photolysis of 2,3,7,8-tetrachlorodibenzo-p-dioxin (2,3,7,8-TCDD) and 1,2,3,4-tetrachlorodibenzo-p-dioxin (1,2,3,4-TCDD). When we exposed the column to a 450 W UV lamp, the adsorbed 1,2,3,4-TCDD or 2,3,7,8-TCDD in 10% 2-propanol/water decomposed completely in 20 minutes and 5 minutes, respectively. The large estimated partition coefficient of 1,2,3,4-TCDD in 10% 2-propanol/water (> 1000) indicates that on the C18 stationary phase, both the saturated hydrocarbon chains and the absorbed 2-propanol may act as proton donors and accelerate the photolysis. In direct sunlight, the adsorbed 1,2,3,4-TCDD in 10% 2-propanol/water decomposed much faster than in a nonaqueous solvent (50% 2-propanol/methanol). This solvent effect is advantageous for the practical use of the C18 photolysis process in aqueous waste treatment. We have demonstrated that complete C18 trapping with continuous photodecomposition of TCDD contained in an aqueous alcohol waste is possible.

The effect of in ovo or day-of-hatch subcutaneous antibiotic administration on competitive exclusion culture (PREEMPT) establishment in neonatal chickens.

The effect of in ovo or day-of-hatch subcutaneous antibiotic administration on the detection of antibiotic residues in yolk sac or blood serum samples and the potential for observed residues to interfere with competitive exclusion (CE) culture establishment was compared in three experiments. The in ovo or subcutaneous administration of gentamicin sulfate or ceftiofur sodium was associated with detectable levels of antibiotic residues in yolk sac or blood serum samples in Experiment 1. Further, the ability to detect antibiotic residues in day-of-hatch chicks was associated with reduced levels of CE culture establishment when cecal propionate level, an indicator of PREEMPT establishment, was determined following PREEMPT application by oral gavage on the day of hatch in Experiments 1 and 2. Restricting chicks from feed, as opposed to providing access ad libitum to a starter ration, for 6 h immediately following administration of PREEMPT improved (P < 0.05) CE culture establishment in Experiment 2 in nonantibiotic injected control chicks, but did not affect (P > 0.05) experimental groups receiving either gentamicin sulfate or ceftiofur sodium by either in ovo or subcutaneous routes. The in ovo administration of 0.1 or 0.2 mg ceftiofur sodium to individual embryos on Day 18 of embryogenesis in Experiment 3 was associated with marked depressions (P < 0.05) in cecal propionate levels compared with uninjected control chicks. When feed was restricted and the time of PREEMPT administration was delayed for 48 or 72 h posthatch, mean cecal propionate levels in in ovo ceftiofur sodium-injected chicks were not significantly different (P > 0.05) from controls, indicating a time- and feed restriction-associated effect on improving CE culture establishment.

Evaluation of potential disinfectants for preslaughter broiler crop decontamination.

The broiler crop has recently been implicated as a major source of Salmonella contamination at commercial processing. Furthermore, feed withdrawal has been positively correlated with increased Salmonella incidence in the crop, probably due to coprophagy. In the present study, a rapid screening assay was developed to evaluate several potential disinfectants in the presence of large quantities of organic matter, simulating the crop environment. An apparent synergistic combination of d-Limonene (DL) and citric acid (CA) was observed when evaluating the potential to eliminate Salmonella in the presence of organic material. A method of encapsulation of DL and CA was developed for voluntary consumption by broilers during feed withdrawal. During an 8-h feed withdrawal individual 8-wk-old broilers voluntarily consumed an average of 21.5 capsules (total of 3.44 g material). When eight capsules were force-administered to Salmonella-challenged 8-wk-old broilers during an abbreviated 4 h feed withdrawal, Salmonella was not recovered using selective enrichment. To evaluate the effect of voluntary capsule consumption, 8-wk-old broilers were challenged with 1 x 108 cfu of Salmonella 5 d prior to an 8 h feed withdrawal. When these broilers were allowed unlimited continuous access to capsules containing DL/CA during an 8 h feed withdrawal, 24.8 capsules per broiler were ingested without affecting Salmonella recovery from crops. When access to capsules containing DL/CA was limited to the final 45 min of an 8 h feed withdrawal in a similar experiment, an average of 22.2 capsules were consumed by each broiler, resulting in a significant decrease in the number of Salmonella-positive crops. Although a number of practical questions and considerations remain, these data suggest that appropriate disinfectants could be administered during preslaughter feed withdrawal for the purpose of reducing foodborne pathogens in crops.

Effect of lactose administration in drinking water prior to and during feed withdrawal on Salmonella recovery from broiler crops and ceca.

Salmonella contamination of the chicken crop has been reported to increase markedly and significantly during feed withdrawal, probably due to coprophagy, and may contribute to carcass contamination at processing. The effect of prolonged lactose administration (2.5%) in the drinking water on the incidence of Salmonella recovery from broiler crops or ceca was evaluated in seven experiments. In these experiments, all or a percentage (providing seeders and contacts) of 7-wk-old broilers were challenged with approximately 1 x 108 cfu Salmonella enteritidis and provided lactose for 5 or 11 d prior to and during an 18 or 24 h feed withdrawal period. A small but significant lactose-mediated reduction in Salmonella contamination of crops was observed in one of two identical experiments with 18 h feed withdrawal. Extending the feed withdrawal period to 24 h did not improve the ability of lactose to affect Salmonella recovery from crops or ceca. Similarly, lactose did not affect Salmonella recovery when the percentage of birds challenged was reduced to 3 out of 16 and Salmonella recovery from crops or ceca of unchallenged, contact broilers was measured. Extending the duration of exposure to 2.5% lactose in the drinking water from 5 to 11 d did not improve the ability of lactose to affect Salmonella recovery. Taken together, these data suggest that provision of 2.5% lactose in the drinking water during the last 5 to 11 d of growout prior to slaughter will not be useful in an integrated Salmonella control program under commercial conditions.

Evaluation of alternative sampling methods for Salmonella critical control point determination at broiler processing.

Several sampling methods have been developed for the detection of Salmonella on broiler carcasses during commercial processing. The present study evaluated three sampling methodologies for sensitivity of Salmonella detection on processed broiler carcasses. Furthermore, the effect of crop removal or chill tank exposure on the frequency of Salmonella recovery was also examined. In two experiments, swab, skin, and carcass rinse samples were compared for sensitivity of Salmonella detection. The results indicated that culture of swabs was less effective (P < 0.05) for Salmonella detection than either skin or carcass rinse samples. No significant (P > 0.05) differences were observed in Salmonella recovery from culture of skin or carcass rinse. In two subsequent experiments, skin and carcass rinse samples were found to be equally sensitive in their ability to detect Salmonella. Additionally, the stages of processing between feather and crop removal were observed to cause significant (P < 0.05) increases in Salmonella recovery within an individual flock. Similar increases (P < 0.05) in Salmonella recovery were also observed following crop removal and immediately following immersion chilling in two separate flocks. These results suggest that culture of skin samples obtained from the thoracic inlet region may be a viable alternative to the traditional whole carcass rinse method for sensitivity of Salmonella detection. Furthermore, these experiments provided some evidence that the majority of Salmonella cross-contamination of carcasses prior to immersion chilling occurred following evisceration, with the chill tank potentially providing a major site for cross-contamination between Salmonella-negative and-positive flocks.

Optimal cooling and warming rates in the preservation of herpes simplex virus (type 2).

Fast rates of cooling and warming were optimal for survival of herpes simplex virus, type 2. Under these conditions cryoprotectants were not necessary for virus preservation.

Titration of human serum antibodies to Toxoplasma gondii with a simple fluorometric assay.

A new technique, FIAX, has been evaluated for the serodiagnosis of toxoplasmosis. It is based on a "dipstick" principle, and a special fluorometer is used to perform the indirect immunofluorescence test. The procedure appears to be simple and rapid and merits consideration as a useful serological test for toxoplasmosis.

Developing a system for assigning individuals into an appropriate residential setting.

With the goal of maximizing the most suitable placement of elderly persons into different types of residential settings, a mathematical system is being developed, using discriminant function analysis, for replicating interdisciplinary clinical team judgments concerning case specific appropriateness for (1) an institutional setting; (2) an apartment in sheltered housing (providing at least the major meal of the day plus housekeeping services); and (3) an apartment in more traditional housing for the elderly. Two discriminant function systems are being developed--the first for separating elderly persons into those needing institutionalization and those requiring a less supervised setting and the second for separting persons judged not to be in need of institutionalization into those requiring either sheltered housing or some other more independent residency situation. While the first function is considered more fully developed at this time, the second is nevertheless useful in that is suggests forces (variables) predictive of such judgments.

Teratogenicity of three polychlorinated dibenzofurans in C57BL/6N mice.

Polychlorinated dibenzofurans (PCDFs) are widespread environmental contaminants which have been detected in human tissues and implicated in several poisoning incidents. Their toxic effects are similar to those observed with other related halogenated aromatic hydrocarbons such as TCDD. The teratogenic effects of three PCDFs, 1,2,3,7,8-pentachlorodibenzofuran (1-PeCDF), 2,3,4,7,8-pentachlorodibenzofuran (4-PeCDF), and 1,2,3,4,7,8-hexachlorodibenzofuran (HCDF), were assessed in C57BL/6N mice. Pregnant mice were exposed on Gestation Days 10-13 to 10 ml corn oil/kg containing PCDFs. The dams were killed on Gestation Day 18 and maternal and fetal toxicity were assessed. All three compounds were highly teratogenic, with very steep and parallel dose-response curves for the two diagnostic indicators of dioxin-like teratogenicity, hydronephrosis, and cleft palate. 4-PeCDF was the most teratogenic with an ED50 of 36 micrograms/kg for cleft palate and 7 micrograms/kg for hydronephrosis. 4-PeCDF was approximately 4 times as potent as 1-PeCDF and 10 times as potent as HCDF. The teratogenic responses occurred at a dose below that where any obvious maternal or fetal toxicity was detected. Thus, these three compounds cause teratogenic responses similar to those seen with TCDD but are only 1/10 to 1/100 as potent.

Reversed-phase liquid chromatography of the twenty-two tetrachlorodibenzo-p-dioxin isomers on pyrenylethyl- and octadecylsilylated silica gel columns.

To purify the tetrachlorobenzo-p-dioxins synthesized for chemical and biological reference standards, reversed-phase liquid chromatography on pyrenylethyl- and octadecylsilylated stationary phase (PYE and C18) columns was employed with 100% methanol. The pyrenylethyl phase satisfactorily separated isomers resulting from mixtures of reaction products which had not been adequately separated with conventional C18 or silica gel. The use of a single chromatographic mode, liquid chromatography, and a single mobile phase with columns of distinctly different properties, C18 and PYE, separates 20 of the 22 isomers from each other and from the 1246/1249 pair. (The observed properties of the two columns afford insight into steric and polarizability differences among isomers.)

Surveillance for Creutzfeldt-Jakob disease among persons with hemophilia.

BACKGROUND: Although Creutzfeldt-Jakob disease (CJD) has been shown to be transmissible through blood components in rodent models, no human blood-to-blood transmission has been documented. If blood transmission were possible in humans, persons with hemophilia in the United States would be at higher risk of contracting CJD, because they receive large numbers of blood components. Nearly one-half of the hemophilia population contracted HIV in the 1980s, and many of these people have since died with neurologic complications. This study investigated whether some hemophilia patients with neurologic disorders may have died with CJD. STUDY DESIGN AND METHODS: Hemophilia treatment Centers across the United States were invited to participate in this retrospective surveillance study. The centers were asked to send any available formalin-fixed paraffin block brain samples from hemophilia decedents. Slides were prepared at the Centers for Disease Control and Prevention and reviewed by three expert neuropathologists. Two slides were stained for the prion protein at the request of one of the neuropathologists. RESULTS: Specimens from 24 decedents with genetic bleeding disorders were collected and reviewed.The panel found no evidence of CJD in any of the specimens. CONCLUSIONS: Although the study sample is small, these results support the growing evidence that CJD is not being transmitted in the nation's blood supply.

Analytical characterization of peptide contaminants of L-tryptophan.

Reversed-phase liquid chromatographic fractions of extracts of 2 preparations of eosinophilia-myalgia syndrome (EMS)-associated L-tryptophan were analyzed by proton nuclear magnetic resonance spectrometry, mass spectrometry, microbial-growth inhibition, and amino acid residue analyses. Fraction components demonstrated properties of an antibiotic peptide resembling bacitracin. Many peptide antibiotics like bacitracin are secondary metabolites of Bacillus species, genus of the tryptophan producer organism for the implicated manufacturer. In order to determine whether a correlation exists between individual EMS cases and the concentration of peptides or bacitracin consumed, reliable methods must be developed for quantification of the total of isoforms.

Genetic factors associated with thrombosis in pregnancy in a United States population.

OBJECTIVE: Polymorphisms in the genes for factor V (factor V Leiden), prothrombin, methylenetetrahydrofolate reductase, and angiotensin-converting enzyme have been associated with the occurrence of venous thrombosis. The objective of this study was to determine the relationships of these polymorphisms to thrombosis during pregnancy. STUDY DESIGN: This case-control study included 41 case patients with venous thrombosis during pregnancy and 76 control subjects matched for hospital and for race (white vs black) who had a normal pregnancy. RESULTS: Among white subjects, mutations in the genes for factor V and prothrombin were associated with increased risks of venous thrombosis during pregnancy (factor V: odds ratio, 18.3; 95% confidence interval, 2.7-432; P =.001; prothrombin: odds ratio infinity; 95% lower confidence limit, 1.7; P =.01). No black subject had either of these two mutations. For both black and white subjects the D/D genotype of the gene for angiotensin-converting enzyme entailed increased risk compared with the other genotypes (odds ratio, 2.7; 95% confidence interval, 1.2-6.3; P =.02). The polymorphism in the gene for methylenetetrahydrofolate reductase was unrelated to thrombosis during pregnancy among both blacks and whites. CONCLUSION: Women who had thrombotic complications during pregnancy demonstrated an increased prevalence of genetic mutations related to coagulation. The additional risk of thrombosis during pregnancy associated with such genetic mutations can be substantial.

Investigation of porcine parvovirus among persons with hemophilia receiving Hyate:C porcine factor VIII concentrate.

BACKGROUND: Porcine clotting factor has been used for more than 15 years to treat severe bleeding episodes in persons with hemophilia who have antibodies to human clotting factor. In 1996, QC procedures revealed for the first time the presence of porcine parvovirus (PPV) in the product. This report describes an investigation to determine the extent of product contamination and to evaluate past recipients of porcine clotting factor (Hyate:C, Speywood Biopharm) for evidence of PPV infection. STUDY DESIGN AND METHODS: Stored specimens from 22 lots of previously released Hyate:C were tested for the presence of PPV DNA by PCR and nested PCR assays. Serum specimens from 98 recipients of Hyate:C and 24 controls who did not receive Hyate:C were tested for PPV antibodies by an immunofluorescence assay. RESULTS: PPV DNA was detected in product from 21 of the 22 lots of Hyate:C, primarily by nested PCR testing. In contrast, none of the serum specimens from the 98 Hyate:C recipients tested positive for PPV IgG antibodies. CONCLUSION: The risk of human disease from percutaneous exposure to low levels of PPV seems to be low. Nevertheless, the theoretical risk of human infection with PPV has led to manufacturing changes, including PCR screening of all porcine plasma, which are designed to eliminate this risk.