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Nitrile hydratases (NHase) catalyze the hydration of nitriles to the corresponding amides. We report on the heterologous expression of various nitrile hydratases. Some of these enzymes have been investigated by others and us before, but sixteen target proteins represent novel sequences. Of 21 target sequences, 4 iron and 16 cobalt containing proteins were functionally expressed from Escherichia coli BL21 (DE3) Gold. Cell free extracts were used for activity profiling and basic characterization of the NHases using the typical NHase substrate methacrylonitrile. Co-type NHases are more tolerant to high pH than Fe-type NHases. A screening for activity on three structurally diverse nitriles was carried out. Two novel Co-dependent NHases from Afipia broomeae and Roseobacter sp. and a new Fe-type NHase from Gordonia hydrophobica were very well expressed and hydrated methacrylonitrile, pyrazine-carbonitrile, and 3-amino-3-( p -toluoyl)propanenitrile. The Co-dependent NHases from Caballeronia jiangsuensis and Microvirga lotononidis , as well as two Fe-dependent NHases from Pseudomonades , were-in addition-able to produce the amide from cinnamonitrile. Summarizing, seven so far uncharacterized NHases are described to be promising biocatalysts.

Background: A comprehensive primary healthcare (PHC) approach requires clear referral and continuity of care pathways. South Africa is a lower-middle income country (LMIC) that lacks data on the role of intermediate care (IC) services in the health system. This study described the model of service provision at one facility in Cape Town, including reason for admission, the mix of services and skills provided and needed, patient satisfaction, patient outcome and articulation with other services across the spectrum of care., Methods: A multi-method design was used. Sixty-eight patients were recruited over one month in mid-2011 in a prospective cohort. Patient data were collected from clinical record review and an interviewer-administered questionnaire, administered shortly after admission to assess primary and secondary diagnosis, referring institution, knowledge of and previous use of home based care (HBC) services, reason for admission and demographics. A telephonic questionnaire at 9-weeks post-discharge recorded their vital status, use of HBC post-discharge and their satisfaction with care received. Staff members completed a self-administered questionnaire to describe demographics and skills. Cox regression was used to identify predictors of survival., Results: Of the 68 participants, 38% and 24% were referred from a secondary and tertiary hospital, respectively. Stroke (35%) was the most common single reason for admission. The three most common reasons reported why care was better at the IC facility were staff attitude, the presence of physiotherapy and the wound care. Even though most patients reported admission to another health facility in the preceding year, only 13 patients (21%) had ever accessed HBC and only 25% (n=15) of discharged patients used HBC post-discharge. Of the 57 patients traced on follow-up, 21(37%) had died. The presence of a Care-plan was significantly associated with a 62% lower risk of death (hazard ratio: 0.38; CI 0.15-0.97). Notably, 46% of staff members reported performing roles that were outside their scope of practice and there was a mismatch between what staff reported doing and their actual tasks., Conclusion: Clients understood this service as a caring environment primarily responsible for rehabilitation services. A Care-plan beyond admission could significantly reduce mortality. There was poor referral to and poor articulation with HBC services. IC services should be recognised as an integral part of the health system and should be accessible., (© 2018 The Author(s); Published by Kerman University of Medical Sciences. This is an open-access article distributed under the terms of the Creative Commons Attribution License (http://creativecommons.org/licenses/by/4.0), which permits unrestricted use, distribution, and reproduction in any medium, provided the original work is properly cited.)

Perfluorinated alkyl acids (PFAAs) are environmental contaminants that have been used in many products for over 50 years. Interest and concern has grown since 2000 on the widespread presence of PFAAs, when it was discovered that PFAAs were present in wildlife samples around the northern hemisphere. Since then, several studies have reported PFAAs in wildlife from many locations, including the remote regions of Antarctica and the Arctic. Although there are a multitude of studies, few have reported PFAA concentrations in reptiles and wildlife in the Southern Hemisphere. This study investigated the presence of PFAAs in the plasma of Nile crocodiles (Crocodylus niloticus) from South Africa. Crocodiles were captured from five sites in and around the Kruger National Park, South Africa, and plasma samples examined for PFAAs. Perfluorooctane sulfonate (PFOS) was the most frequent PFAA detected; with median values of 13.5 ng/g wet mass in crocodiles. In addition to PFOS, long chain perfluorinated carboxylic acids were also detected. Correlations between total length and PFAA load were investigated, as were differences in PFAA accumulation between sexes. No correlations were seen between crocodile size, nor were there sex-related differences. Spatial differences were examined and significant differences were observed in samples collected from the different sites (p < 0.05). Flag Boshielo Dam had the highest PFOS measurements, with a median concentration of 50.3 ng/g wet mass, when compared to the other sites (median concentrations at other sites below 14.0 ng/g wet mass). This suggests a point source of PFOS in this area., (Published by Elsevier Ltd.)

Introduction: Although antiretroviral therapy (ART) reduces individual tuberculosis (TB) risk by two-thirds, the population-level impact remains uncertain. Cape Town reports high TB notification rates associated with endemic HIV. We examined population trends in TB notification rates during a 10-year period of expanding ART., Methods: Annual Cape Town TB notifications were used as numerators and mid-year Cape Town populations as denominators. HIV-stratified population was calculated using overall HIV prevalence estimates from the Actuarial Society of South Africa AIDS and Demographic model. ART provision numbers from Western Cape government reports were used to calculate overall ART coverage. We calculated rates per 100,000 population over time, overall and stratified by HIV status. Rates per 100,000 total population were also calculated by ART use at treatment initiation. Absolute numbers of notifications were compared by age and sub-district. Changes over time were described related to ART provision in the city as a whole (ART coverage) and by sub-district (numbers on ART)., Results: From 2003 to 2013, Cape Town's population grew from 3.1 to 3.7 million inhabitants, and estimated HIV prevalence increased from 3.6 to 5.2%. ART coverage increased from 0 to 63% in 2013. TB notification rates declined by 16% (95% confidence interval (CI), 14-17%) from a 2008 peak (851/100,000) to a 2013 nadir (713/100,000). Decreases were higher among the HIV-positive (21% (95% CI, 19-23%)) than the HIV-negative (9% (95% CI, 7-11%)) population. The number of HIV-positive TB notifications decreased mainly among 0- to 4- and 20- to 34-year-olds. Total population rates on ART at TB treatment initiation increased over time but levelled off in 2013. Overall median CD4 counts increased from 146 cells/µl (interquartile range (IQR), 66, 264) to 178 cells/µl (IQR 75, 330; p<0.001). Sub-district antenatal HIV seroprevalence differed (10-33%) as did numbers on ART (9-29 thousand). Across sub-districts, infant HIV-positive TB decreased consistently whereas adult decreases varied., Conclusions: HIV-positive TB notification rates declined during a period of rapid scale-up of ART. Nevertheless, both HIV-positive and HIV-negative TB notification rates remained very high. Decreases among HIV positives were likely blunted by TB remaining a major entry to the ART programme and occurring after delayed ART initiation.

The onslaught on the World's rhinoceroses continues despite numerous initiatives aimed at curbing it. When losses due to poaching exceed birth rates, declining rhino populations result. We used previously published estimates and growth rates for black rhinos (2008) and white rhinos (2010) together with known poaching trends at the time to predict population sizes and poaching rates in Kruger National Park, South Africa for 2013. Kruger is a stronghold for the south-eastern black rhino and southern white rhino. Counting rhinos on 878 blocks 3x3 km in size using helicopters, estimating availability bias and collating observer and detectability biases allowed estimates using the Jolly's estimator. The exponential escalation in number of rhinos poached per day appears to have slowed. The black rhino estimate of 414 individuals (95% confidence interval: 343-487) was lower than the predicted 835 individuals (95% CI: 754-956). The white rhino estimate of 8,968 individuals (95% CI: 8,394-9,564) overlapped with the predicted 9,417 individuals (95% CI: 7,698-11,183). Density- and rainfall-dependent responses in birth- and death rates of white rhinos provide opportunities to offset anticipated poaching effects through removals of rhinos from high density areas to increase birth and survival rates. Biological management of rhinos, however, need complimentary management of the poaching threat as present poaching trends predict detectable declines in white rhino abundances by 2018. Strategic responses such as anti-poaching that protect supply from illegal harvesting, reducing demand, and increasing supply commonly require crime network disruption as a first step complimented by providing options for alternative economies in areas abutting protected areas.

Repeated annual episodes of Nile crocodile deaths in two isolated areas of the Kruger National Park prompted the investigation of possible organohalogen pollutant involvement. Crocodile eggs were collected close to one of the mortality sites (Gorge) as well as from a crocodile farm (CF) as reference. ∑DDT was significantly higher in Gorge (450ng/g wm) than in CF eggs (85ng/g wet mass). Percentage DDT of ∑DDT was significantly higher in CF (14 per cent) than in Gorge eggs (5 per cent). Mean ∑DDT was almost 70 times higher than mean ∑PCB in Gorge eggs. HCB, β-HCH, mirex, brominated flame retardants (BFRs), and perfluorinated compounds (PFCs) occurred at lower concentrations. We believe that the BFR and PFCs data represent the first published results for any crocodile egg. Thickening of the outer eggshell layer of Gorge eggs was significantly associated with higher concentrations of ∑DDT. Concentrations of ∑DDT and other pollutants were in the same range as eggs from elsewhere, where there were no mortalities. Concentrations of ∑DDT in eggs from healthy Australian crocodiles were of the same orders of magnitude as the current study, making it highly unlikely that the concentrations of pollutants measured in the present study would have caused or substantially contributed towards the mortalities observed. Concerns about reproduction and behaviour remain. As large predators, crocodilians are at the apex of the freshwater aquatic food web. More research is needed to guide measures to manage African freshwater systems so that it will also sustainably accommodate these large, long-lived animals., (Copyright © 2013 Elsevier Inc. All rights reserved.)

Background: Antiretroviral therapy (ART) in pregnancy is a crucial intervention in the prevention of mother-to-child transmission (PMTCT) of HIV. It is recognised that mother-to-child transmission is reduced with each week on ART. However, in most South African settings, ART initiation is delayed owing to slow determination of treatment eligibility and separation of HIV and antenatal care services., Objective: The rapid initiation of an ART in pregnancy programme is a model of care designed to expedite treatment initiation in ART- eligible pregnant women. This study evaluated the performance of the programme., Methods: Participants enrolled in the ART programme in the same week as their first ANC visit throughout 2011, and had outcome data available by March 2012. Treatment eligibility was determined or confirmed via point-of-care CD4+ testing. Eligible women were offered ART immediately, with concurrent counselling and safety laboratory blood testing. Women attended until 6 - 8 weeks after delivery. Data were collected from clinical records with infant polymerase chain reaction (PCR) results at 6 weeks., Results: Of 134 ART-eligible (CD4+ count <350 cells/μl or WHO stage III/IV) pregnant women, 130 (97.0%) started ART, 118 (90.8%) initiating treatment the same day that treatment eligibility was determined. There were no abnormal laboratory blood results or adverse events that required medical intervention. Pre-delivery retention in care and infant mortality were comparable to those in similar settings. Of the 107 pregnancies with PCR outcomes available, there was 1 positive HIV result in an infant (0.9%). Maternal viral load suppression in this mother was not achieved by the time of delivery., Conclusions: This pilot programme provides evidence that rapid ART initiation in pregnancy is safe, feasible and effective in reducing PMTCT. Further follow-up is required to monitor long-term outcomes.

Objective: To describe the nature of hearing loss and associated risk profile in a South African population of infants and children diagnosed at a pediatric referral clinic., Methods: A retrospective review of patient files for a pediatric auditory evoked potential clinic in Pretoria was conducted (January 2007-December 2011). Collected data included demographical information, risk factors from case history questionnaire, diagnosis (type and degree of hearing loss), documented age of caregiver suspicion and age of first diagnosis., Results: Hearing loss was present in 73% (73/100) of cases evaluated. Permanent hearing losses (SNHL, ANSD and mixed) constituted 76% of losses. Unilateral hearing losses constituted 8% of SNHL and 20% of conductive hearing loss. ANSD was diagnosed in 21.4% and SNHL in 78.6% of permanent non-conductive hearing loss cases. The most prevalent SNHL risk was family history of hearing loss and for ANSD it was admittance to the NICU for more than 5 days. The majority of the sample was diagnosed with a permanent bilateral SNHL and ANSD after 36 months of age (47%) despite 40% already suspected of having a hearing loss before 12 months of age., Conclusions: A high prevalence of ANSD was found with preventable risk factors often indicated. Age of diagnosis was significantly delayed, evidencing the lack of early hearing detection services in South Africa. The majority of children were diagnosed at ages precluding optimal benefits from early detection and subsequent intervention., (Copyright © 2012 Elsevier Ireland Ltd. All rights reserved.)

Objective: To investigate patient education and counseling activities prior to the initiation of antiretroviral therapy (ART) at public sector services across Cape Town, South Africa., Methods: Key informant interviews and programme reviews were conducted with government bodies and non-governmental organisations involved in patient preparation activities., Results: All 11 organisations in Cape Town involved in training and managing personnel to prepare patients for ART during 2010 participated. Each organisation reported a different approach to patient preparation within public sector clinics and in each aspect of patient preparation activities. The number of patient education sessions ranged from 3 to 7, and the delays to ART initiation introduced by patient preparation ranged from 3 to 6 weeks. Different patient education materials (pamphlets, posters and flipcharts) were used by various programmes, and all programmes reported that shortages in materials meant that patient preparation often took place without any educational materials. Each programme also reported attention to mental illness and alcohol/substance use disorders, but none employed formal screening tools consistently, and the handling of patients with potential mental health- or substance-related problems varied., Conclusion: Approaches to prepare patients before ART initiation are wide ranging in one part of South Africa. Their relative value requires investigation, as there is little evidence for the impact of varying approaches. Moreover, the risks associated with delayed ART initiation may outweigh any benefits of patient education before the start of treatment., (© 2012 Blackwell Publishing Ltd.)

Initiation of antiretroviral therapy (ART) in pregnancy is an important intervention to prevent the mother-to-child transmission (MTCT) of HIV and to promote maternal health. Early initiation of ART is particularly important to achieve viral suppression rapidly before delivery. However, current approaches to start ART in pregnancy may be problematic in many settings, with referrals between antenatal care (ANC) and ART services, and delays for patient preparation before ART initiation. These steps contribute to a sizable proportion of women failing to receive adequate duration of ART before delivery, increasing the risk of MTCT. To address these limitations, we developed the rapid initiation of antiretroviral therapy in pregnancy (RAP) programme. The programme featured the use of point-of-care CD4 testing to identify ART-eligible women with CD4 cell counts ≤ 350 cells/µl; immediate ART initiation in women on the same day that eligibility was determined, if possible; and intensive counselling and support for ART initiation during the first few weeks on ART. We implemented RAP in an antenatal clinic setting in Cape Town South Africa. Between February and August 2011, a total of 221 HIV-infected women were referred to the programme for CD4 cell count testing and 101 (46%) were eligible for ART. Of these, 98 women (97%) started therapy during pregnancy, 89 (91%) on the day of referral to the service. In-depth interviews suggested that although there were substantial challenges facing HIV-infected women initiating ART in pregnancy, the availability of immediate services and counselling support played an important role in addressing these. While further research is needed, this evaluation demonstrates that a novel service delivery approach may facilitate rapid ART initiation in pregnancy.

Erythema nodosum (EN) may follow a variety of infections, but in regions with a high prevalence of tuberculosis, is frequently associated with a positive tuberculin skin test (TST) and tuberculosis infection. We aimed to investigate the immunological differences between patients with EN as a manifestation of primary tuberculosis, and those with progressive pulmonary tuberculosis (PTB) or asymptomatic infection. We studied the inflammatory response to both mycobacterial and non-mycobacterial antigens in 11 children with EN associated with a positive TST, 22 children with culture-confirmed tuberculosis, and 53 healthy skin test-positive children. In addition, we evaluated functional anti-mycobacterial immunity using an ex vivo assay of mycobacterial growth restriction in five children with EN and 15 with PTB. Patients with EN were distinguished by enhanced mycobacterial growth restriction on the functional assay, which was associated with a markedly increased production of IFNgamma in response to stimulation with purified protein derivative of Mycobacterium tuberculosis. Children presenting with EN and a positive TST show evidence of responses associated with enhanced anti-mycobacterial immunity.