Prevalence of vaccine-derived polioviruses in stools of immunodeficient children in South Africa.

AIMS: The aim of the study was to determine the prevalence of vaccine-derived polioviruses (VDPVs) in stool specimens of immunodeficient patients such as HIV-positive children (including those with an AIDS indicator condition, according to the Centres for Disease Control and Prevention classification) by applying various molecular techniques. METHODS AND RESULTS: A total of 164 stool samples from HIV-positive children and 23 stool samples from healthy immunocompetent children (the control group) were analysed during 2003 and 2004. By applying a reverse transcription polymerase chain reaction (RT-PCR) in combination with a nested PCR, a total of 54 enteroviruses were detected in the stool specimens of the immunodeficient children. The use of restriction enzymes and a Sabin specific RT-triplex PCR confirmed the presence of 13 polioviruses (PVs), such as seven Sabin PV type 1, four Sabin PV type 3 and two Sabin PV type 2 isolates. The 5'untranslated region and the VP1 capsid-encoding protein of the 13 PVs and the three PVs from the stools of the immunocompetent children were partially sequenced and their genetic relatedness was deduced from the constructed phylogenetic trees. The majority of the PVs isolated from the stools of the immunodeficient children (10 of 13 isolates) were classified as 'oral poliovirus vaccine (OPV)-like viruses', as these isolates had close sequence relationships (>99% in VP1 nucleotide sequences) to the original Sabin PV vaccine strains. Three PVs showed < or =99% VP1 sequence identity to the Sabin PV vaccine strains and were classified as 'suspected' immunodeficient VDPVs (iVDPVs). All of the OPV-like isolates and the 'suspected' iVDPVs carried mutations at specific positions in their partially sequenced regions, which have been associated with reversion of the attenuated Sabin PV vaccine strains to increased neurovirulence. CONCLUSIONS: Thus, this study adds further evidence to the observation that immunodeficient individuals may excrete OPV strains with potential neurovirulent phenotypes. SIGNIFICANCE AND IMPACT OF THE STUDY: Prolonged excretion of PVs by immunodeficient individuals is of major concern, because continued replication of PVs in the human gut could result in the reversion of these viruses to greater neurovirulence. When exposed to OPV, immunodeficient patients may become chronically infected, spreading potentially neurovirulent VDPVs for many months or years to close contacts and children who are no longer being vaccinated after termination of OPV vaccination in the near future.

Poliovirus vaccine strains in sewage and river water in South Africa.

Since the initiation of the global poliomyelitis eradication program in 1988, the number of wild-type polio cases decreased from 350,000 to fewer than 500, and the number of polio endemic countries declined from more than 125 to 10. The last case of polio in South Africa caused by a wild-type poliovirus (PV) occurred in 1989. The live attenuated oral poliovirus vaccine (OPV) has been effectively used in the reduction and control of poliomyelitis. However, as OPV strains are excreted in stools after vaccination, this vaccine could become a source of dissemination of PVs in the environment and the potential cause of poliomyelitis. Therefore, the aim of the study was to determine the occurrence of OPV strains in selected sewage and river water samples. During the period between 2001 and 2003, 138 samples of river water and 213 samples of settled sewage were collected from selected areas of South Africa. A total of 860 plaques were analysed, which consisted of 703 plaques from the sewage and 157 plaques from the river water samples. Using a reverse transcriptase (RT)-multiplex PCR, 49 PVs were successfully distinguished from 176 non-polio enteroviruses (NPEVs). The 176 NPEVs consisted of 50 coxsackie B2 viruses (CBV2), followed by 39 echoviruses 11 (ECV11), 25 CBV5, 21 CBV3, 15 CBV4, 14 coxsackie A6 viruses (CAV6), 7 CBV6, 2 CAV5, 2 CBV1, and 1 ECV19, which was in agreement with the prevalence of these EVs in other parts of the world. The Sabin-specific RT-triplex PCR revealed the presence of 29 Sabin PV type 1, 8 Sabin PV type 2, and 12 Sabin PV type 3 isolates. Buffalo green monkey kidney and primary liver carcinoma cell cultures allowed the amplification of a broad spectrum of EVs, whereas human epidermoid carcinoma cells were more selective for PVs. This study addressed some of the issues regarding the prevalence of OPV strains in the environment. The identification of 49 viable OPV isolates confirmed the presence and circulation of PV vaccine strains in sewage and river water. The extent of the potential health risk constituted by these OPV isolates remains to be investigated.

Prevalence of vaccine-derived polioviruses in sewage and river water in South Africa.

Polioviruses (PVs) are not associated with waterborne transmission to the same extent as many other enteric viruses. However, they are typically transmitted by the faecal-oral route, which implies that the risk of infection by exposure to the viruses in water cannot be underestimated. The risk appears particularly high for rural communities, which use sewage-polluted river water for domestic purposes. Thus, the presence in the environment of highly evolved, neurovirulent vaccine-derived poliovirus (VDPV) strains in the absence of polio cases would have important implications for strategies to terminate immunisation with oral poliovirus vaccine (OPV) following global polio eradication. The aim of the current study was to determine the prevalence of VDPVs in selected sewage and river water samples collected from 2001 to 2003, and to construct phylogenetic trees of the partially sequenced 5'untranslated region (5'UTR) and the VP1 region of the genomes to deduce the genetic relatedness between the PV strains. Using the monolayer plaque assay, 703 plaques from sewage and 157 plaques from river water samples were analysed. Application of a RT-multiplex PCR revealed that 176 of these plaques were non-polio enteroviruses, and 49 were PV isolates. The Sabin-specific RT-triplex PCR revealed the presence of 29 Sabin PV type 1, 8 Sabin PV type 2 and 12 Sabin PV type 3 isolates. The 5'UTR and the VP1 region of 13 PV type 1, 7 PV type 3 and 6 PV type 2 isolates were partially sequenced. The majority of the OPV isolates (24 out of 26) displayed close sequence relationships (>99% VP1 sequence identity) to the parental Sabin PV vaccine strains and were classified as "OPV-like viruses". Two isolates (D1 08/28 and OF1 05/21) were found to be highly divergent and were classified as "suspected" VDPVs. Isolate OF1 05/21 (a "suspected" VDPV type 1) showed more than 0.9% divergence in VP1, whereas isolate D1 08/28 (a "suspected" VDPV type 2) showed 1.4% divergence in VP1 from the parental Sabin PV vaccine strains. As with most of the other OPV-like isolates, these "suspected" VDPVs were carrying mutations, which have previously been associated with reversion of the attenuated Sabin PV strains to increased neurovirulence. It was estimated that the total period of replication for the two "suspected" VDPVs was between 12 and 16 months. In conclusion, this study provided new and relevant information on the prevalence of "suspected" VDPVs in sewage and river water, and opened the way to assess the possible broader significance of the findings reported here.

Detection of enteroviruses in untreated and treated drinking water supplies in South Africa.

Enteric viruses have been detected in many drinking water supplies all over the world. A meaningful number of these supplies were treated and disinfected according to internationally acceptable methods. In addition, counts of bacterial indicators (coliform bacteria and heterotrophic plate count organisms) in these water supplies were within limits generally recommended for treated drinking water and these findings have been supported by epidemiological data on infections associated with drinking water. The shortcomings of conventional treatment methods and indicator organisms to confirm the absence of enteric viruses from drinking water, was generally ascribed to the exceptional resistance of these viruses. In this study, the prevalence of enteroviruses detected from July 2000 to June 2002 in sewage, river-, borehole-, spring- and dam water as well as drinking water supplies treated and disinfected according to international specifications for the production of safe drinking water was analysed. A glass wool adsorption-elution technique was used to recover viruses from 10--20 l of sewage as well as environmental water samples, in the case of drinking water from more than 100 l. Recovered enteroviruses were inoculated onto two cell culture types (BGM and PLC/PRF/5 cells) for amplification of viral RNA with nested-PCR being used to detect the amplified viral RNA. Results from the study demonstrated the presence of enteroviruses in 42.5% of sewage and in 18.7% of treated drinking water samples. Furthermore, enteroviruses were detected in 28.5% of river water, in 26.7% of dam/spring water and in 25.3% of borehole water samples. The high prevalence of coxsackie B viruses found in this study suggested, that a potential health risk and a burden of disease constituted by these viruses might be meaningful. These findings indicated that strategies, other than end-point analysis of treated and disinfected drinking water supplies, may be required to ensure the production of drinking water that does not exceed acceptable health risks. More reliable approaches to ensure acceptable safety of drinking water supplies may be based on control by multiple-barrier principles from catchment to tap using hazard assessment and critical control point (HACCP) principles.

[Stimulation of adrenocortical secretion in healthy persons with hypoglycemia provoked by insulin]. / Stimulirane na adreno-kortikalnata sekretsiia u zdravi litsa s khipoglikemiia, predizvikana ot insulin

Results are reported from the carried out test for adrenocortical secretion stimulation with hypoglycemia, provoked by insulin in 63 healthy individuals. Plasma cortisol elevates to 37,4 mkg from the average initial levels 16,1 mkg/100 ml. The elevation is with 21,3 mkg/100 ml. The indices for the negative (abnormal) hormonal response are derived--when the elevation is under 5,4 mkg and the plasma cortisol level after stimulation with insulin remains under 19,7 mkg/100 ml. The clinical application of ITT is discussed.