Novel PFAS-specific monitoring approach for highly impacted surface waters.

In regulatory environmental monitoring programs, only a very small fraction of the vast number of per- and polyfluoroalkyl substances (PFAS) are investigated by target analysis. Therefore, non-target analysis (NTA) studies are increasingly conducted to detect unknown or unnoticed PFAS. These studies are often based on a few grab samples. Thus, discontinuously emitted PFAS from industrial batch processes might be easily overlooked. To address this deficiency and obtain in-depth information on the occurrence and temporal trend of PFAS in surface water impacted by treated industrial waste water, a comprehensive target and NTA study was implemented for 29 months. Elevated PFAS concentrations with up to 10.8 µg L-1 were detected in the river water by target analysis. In addition to PFAS target analysis, the water samples were analyzed by liquid chromatography-high-resolution tandem mass spectrometry (LC-HRMS/MS). Data processing strategies and various filtering steps were applied to prioritize PFAS. Substances were identified by comparing data to available internal and external PFAS suspect lists, a fragment ion and neutral loss list, and spectral libraries. Several compounds were unequivocally identified based on reference standards. Fifty-five PFAS were (tentatively) identified using NTA. Of those, 43 could be assigned to 13 different homologous series. Partly fluorinated short-chain carboxylic acids (H-PFCA) and sulfonic acids (H-PFSA) were predominantly found in addition to perfluoroalkyl carboxylic acids (PFCA) and the alkyl ether carboxylic acid DONA. To the best of our knowledge, 12 PFAS were reported in surface water for the first time. Signal intensities of individual PFAS and signal ratios varied widely over time, which may indicate batch operations leading to discontinuous emission. Results and insights from this screening approach on PFAS can be used to optimize forthcoming surface water monitoring programs by including newly identified PFAS and selecting appropriate sampling intervals.

Quality assessment of digested sludges produced by advanced stabilization processes.

The European Union (EU) Project Routes aimed to discover new routes in sludge stabilization treatments leading to high-quality digested sludge, suitable for land application. In order to investigate the impact of different enhanced sludge stabilization processes such as (a) thermophilic digestion integrated with thermal hydrolysis pretreatment (TT), (b) sonication before mesophilic/thermophilic digestion (UMT), and (c) sequential anaerobic/aerobic digestion (AA) on digested sludge quality, a broad class of conventional and emerging organic micropollutants as well as ecotoxicity was analyzed, extending the assessment beyond the parameters typically considered (i.e., stability index and heavy metals). The stability index was improved by adding aerobic posttreatment or by operating dual-stage process but not by pretreatment integration. Filterability was worsened by thermophilic digestion, either alone (TT) or coupled with mesophilic digestion (UMT). The concentrations of heavy metals, present in ranking order Zn > Cu > Pb > Cr ~ Ni > Cd > Hg, were always below the current legal requirements for use on land and were not removed during the processes. Removals of conventional and emerging organic pollutants were greatly enhanced by performing double-stage digestion (UMT and AA treatment) compared to a single-stage process as TT; the same trend was found as regards toxicity reduction. Overall, all the digested sludges exhibited toxicity to the soil bacterium Arthrobacter globiformis at concentrations about factor 100 higher than the usual application rate of sludge to soil in Europe. For earthworms, a safety margin of factor 30 was generally achieved for all the digested samples.

[Not Available]. / Caliciviren Virale Auslöser akuter Gastroenteritiden : Virale Auslöser akuter Gastroenteritiden.

Human caliciviruses are highly infectious and co-circulate worldwide. They are responsible for many individual cases and extensive outbreaks of acute gastroenteritis. The ability of the viruses to survive in the environment facilitates the fecal-oral spread through water, food, aerosols, and person-to-person contact. Therefore, they are an important global public health problem. Despite the lack of a cell culture or animal model system to grow caliciviruses, great advances in our understanding of these pathogens, especially of the NLVs, have been made by using molecular methods. More and more information about the viral genome is being accumulated in data bases. This information will be used to develop sensitive molecular methods for the better understanding of the viral biology and epidemiology and will assist in developing strategies to prevent and control infections.

Molecular typing of echovirus serotype 4 isolates.

We report on clinical samples Stuttgart/97, Berlin/99 and Jasi/99 associated with aseptic meningitis. All three samples contained echovirus 4 (E4) but Stuttgart/97 was simultaneous infected with echovirus 30 (E30). The genetic relationship of the E4 strains was assessed using RT-PCR and direct sequencing of amplicons derived from the genomic region encoding the capsid protein VP1. The sequences have been compared with each other and with sequences of further E4 strains obtained from GenBank. The analysis confirms that sequences of recent isolates have drifted away from elderly strains over a longer period of time. Several amino acid changes in assumed antigenic sites of the VP1 gene may be sufficient to cause changes in antigenic specificity and therefore they may be a reason for failure of serological typing of some new antigenic E4 variants.

Genetic variability within the VP1 coding region of echovirus type 30 isolates.

Genetic relationships of the prototype Bastianni strain of 1958 and of 13 echovirus type 30 (ECV30) isolates associated with meningitis cases in Germany during a period from 1966 to 1997 were investigated using direct sequencing of amplicons derived from a part of the capsid protein VP1 gene. Sequences were aligned both with each other and with known sequences of other type 30 echovirus strains. Phylogenetic analysis indicated that isolates investigated in this work fell into at least three genetic clusters apart from the prototype Bastianni strain. This suggests that genetically distinct groups of ECV30 variants have developed over time.

Molecular epidemiology of outbreaks of gastroenteritis associated with small round structured viruses in Germany in 1997/98.

The molecular epidemiology of small round structured viruses (SRSVs) in Germany was studied using fecal specimens from 16 SRSV-associated gastroenteritis outbreaks in different parts of Germany during 1997/1998 by reverse transcription-polymerase chain reaction (RT-PCR) and by sequencing of the ORF1 and ORF3 amplicons. The majority of the isolates clustered in one subtype and were closely related to published SRSV sequences of genogroup II.

Characterization of echovirus 25 (ECV 25) in the VP1/2A gene junction region. Brief report.

Genetic relationships among echovirus type 25 (ECV 25) isolates associated with aseptic meningitis in Germany 1997/98 and a 40-year-old ECV 25 prototype strain were investigated using RT-PCR and direct sequencing the VP1/2A gene junction region. Sequences were compared to each other and to non-polio enterovirus representatives (phylogenetic analysis). The analysis indicated that the sequences of recent isolates have drifted over time distinctly away from the prototype strain sequence. Genetic drift may change biological features of isolates possibly leading to new antigenic variants.

Quality control study on the performance of GB virus C/hepatitis G virus PCR.

BACKGROUND/AIMS: A novel virus, GBV-C/HGV, with a genome RNA organization similar to that of the Flaviviridae family was identified in sera of patients with hepatitis. The presence of GBV-C/HGV RNA can only be determined by the amplification of genomic regions using the reverse transcriptase-polymerase chain reaction (RT-PCR). METHODS: To assess the quality of the RT-PCR, 14 laboratories investigated a coded serum panel that comprised three GBV-C/HGV RNA-positive sera from three different patients, dilutions of these sera, and three GBV-C/HGV RNA-negative serum samples, two of which were collected from patients with hepatitis C but without GBV-C/HGV infection. In-house RT-PCR as well as commercially available GBV-C/HGV test kits were used in this study. RESULTS: Only four laboratories (29%) reported the expected results, and four laboratories (29%) false-positive results; nine laboratories (64%) reported at least one false-negative result. Eleven laboratories (79%) detected the undiluted samples. The majority of false results were obtained with the dilutions of GBV-C/HGV RNA-positive samples. Negative results in the 10(-4) dilution were not considered to be false-negative, since during pre-screening GBV-C/HGV RNA had been detected in this dilution in only 50% of assays by the three laboratories involved in organizing the evaluation. Results obtained by commercial kits and by in-house assays were indiscriminate in quality of performance in this study. CONCLUSION: To facilitate further quality assessment studies on the performance of GBV-C/HGV RNA detection, an international GBV-C/HGV RNA standard should be made available. Further efforts are required to optimize GBV-C/HGV RT-PCR.

Epidemiological and clinical aspects of hepatitis G virus infection in blood donors and immunocompromised recipients of HGV-contaminated blood.

BACKGROUND AND OBJECTIVES: The infectiousness and clinical relevance of the newly discovered blood-borne Flaviviridae-like agent, termed hepatitis G virus (HGV), are not well understood. MATERIALS AND METHODS: Twenty-three transfusion recipients of two HGV-affected long-term blood donors were studied for HGV genome and antibodies to the putative envelope 2 glycoprotein (anti-E2) of HGV. Nine recipients had nonhematological disorders and 14 suffered from severe hematological diseases and 7 of them received allogeneic bone marrow or blood stem cell transplantation. The molecular epidemiology of the observed HGV infection was studied by direct sequencing of parts of the 5'-noncoding region, NS3, and NS5 region of HGV in the 2 long-term donors and in their 6 recipients who became HGV RNA positive. Additionally, 549 individuals-homologous (n = 254) and autologous blood donors (n = 202), and medical staff (n = 89)--were investigated for the presence of HGV RNA. RESULTS: HGV RNA in serum was found in 15 of the 23 (65%) transfusion recipients with known exposure of HGV-contaminated blood. Seven of the remaining 8 recipients showed only an anti-E2 response, indicating previous HGV infection with spontaneous clearance of the virus. In one recipient neither HGV RNA nor anti-E2 could be detected. Molecular evidence for HGV transmission by the 2 donors was found in 3 of the 6 recipients studied. The alanine aminotransferase levels were not significantly different in the HGV RNA positive and negative recipients, and none of the 23 recipients developed posttransfusion hepatitis. Persistent HGV infection was observed especially in recipients with severe hematological disorders or in those in whom intensive immunosuppressive treatment was necessary. Of the 549 individuals studied, 10 (1.8%) were healthy carriers of HGV RNA. CONCLUSION: The persistence of transfusion-acquired HGV infection is not associated with acute or chronic hepatitis, but may be influenced by the recipient's underlying disease.

Hepatitis GBV-C sequences in patients infected with HCV contaminated anti-D immunoglobulin and among i.v. drug users in Germany.

BACKGROUND/AIMS: A novel virus, GB virus-C (GBV-C), with a genome organization similar to those of the Flaviviridae family was identified in sera of patients diagnosed with hepatitis. Up to now little has been known about the prevalence of GBV-C sequences in German hepatitis C virus infected patients. METHODS: We investigated two groups of patients chronically infected with hepatitis C virus: (i) Women infected in 1978/79 with HCV-contaminated anti-D immunoglobulin batches in former Eastern Germany, and (ii) i.v. drug users infected with different HCV subtypes. Nested polymerase chain reaction products amplified with GBV-C specific primer pairs within the helicase region were sequenced directly and compared with the GBV-C sequences reported recently. RESULTS: GBV-C sequences of the putative NS 3 gene region were shown to occur in two randomly selected anti-D immunoglobulin batches of 1978 (GI and GII) and two sera of HCV-infected women drawn in 1979. In patient 3, the HCV-infected erythrocyte donor in 1978, specific GBV-C sequences were also evident in serum drawn in 1990. In the high-risk group of i.v. drug users, 49% were GBV-C RNA positive. Among the 21 GBV-C positive samples, 11 were coinfected with HCV subtype 3a and 10 with subtype 1 b. All isolates showed an overall homology to the GBV-C sequence reported by Simons of 75-81% at the nucleotide level and 94-100% at the amino acid level. CONCLUSION: GBV-C sequences are detectable in the anti-D immunoglobulin batches which caused a hepatitis C virus outbreak in 1979, and a first hint of its transmission to recipients was shown. The detection of GBV-C in patient serum drawn 12 years after the onset of chronic liver disease confirms the persistence of the novel virus described here. In the group of i.v. drug users a high frequency of GBV-C sequences (49%) was shown, and the considerable variability of the nucleotide sequences indicates the existence of different GBV-C genotypes/subtypes.

Vaccine-associated cases of poliomyelitis over a 30 year period in East Germany.

A report is presented about studies on poliovirus type 3 isolates from vaccine-associated cases or contacts of cases of paralytic poliomyelitis, observed over a period of 30 years in East Germany (former GDR). In the viral isolates, some mutations were found in comparison to the Sabin vaccine type 3 strain, distributed over the whole genome. The significance of these mutations has been discussed, especially the mutation at position 472 in the 5' noncoding region found in all the isolates investigated. In five isolates, intertypic recombination between Sabin type 3 and Sabin type 1 vaccine strain occurred. Primary and secondary structures were analysed for the recombination sites.

Differentiation of vaccine and wild mumps viruses by polymerase chain reaction and nucleotide sequencing of the SH gene: brief report.

The nucleotide sequences of the SH gene and its F gene flanking region were determined over a range of 322 nucleotides for two live vaccines, two vaccine-associated isolates, six wild mumps viruses, and an attenuated mumps virus and compared to other sequences already published. Comparison revealed that the vaccine strains were clearly different from each other and the postvaccination isolates were different from the vaccines used. The viruses were assigned to three known cocirculating viral lineages. The attenuated mumps virus possesses nucleotide sequences identical to those of its progenitor strain.

Molecular characterization of mumps virus strains circulating during an epidemic in eastern Switzerland 1992/93.

Ten mumps virus isolates recovered during an epidemic in Switzerland between Autumn 1992 and Spring 1993 were compared by nucleotide sequence analysis of the SH gene. The isolates were found to belong to two distinct yet closely related wild type strains in a newly identified European mumps virus lineage.

Complete nucleotide sequence of the neuraminidase gene of the human influenza virus A/Chile/1/83 (H1N1). Brief report.

The complete nucleotide sequence of the neuraminidase (NA) gene of influenza virus A/Chile/1/83 (H1N1) has been determined after reverse transcription and cloning into the plasmid pAT 153/PvuII/8. The gene is 1461 nucleotides long and codes for a protein of 470 amino acids. The overall nucleotide and predicted amino acid sequence of the A/Chile/1/83 NA exhibits a high homology with other N1 neuraminidases. Hyper-variable regions concerning A to G exchanges are discussed.