Low resource innovations for sustaining service delivery: examples from districts in Malawi

Low-income countries have less funds to allocate for water, sanitation & hygiene (WASH) services, but those funds which are available should still be used to have the greatest service delivery reach possible. This paper shares examples from several District Water Development Offices in Malawi of innovations developed to sustain service delivery with the limited resources they do have. These approaches include using available funds to pay for lower-cost alternatives which achieve their objectives, using other networks existing in communities to increase their reach, and leveraging other sets of resources that exist in the area

Mutual reinforcement: combining project outputs with capacity development outcomes for service delivery

Capacity development of permanent local institutions is needed to improve the sustainability of investments made in the water, sanitation and hygiene (WASH) sector. To check capacity development intentions, development partners (DPs) can ask the question “What capacities are you developing and why?” This will verify that capacity development is being done with precise objectives, and is aligned with institutional needs and role definitions. DPs can use implementation and capacity development objectives as mutually reinforcing opportunities to support strong project outputs as well as to improve outcomes for service delivery. Two particular techniques for capitalizing on this duality are highlighted: supporting implementers, and supporting reflective learning. Examples of practical combinations of capacity development approaches are presented from the perspective of Engineers Without Borders Canada working in collaboration with other DPs and with district governments in Malawi’s WASH sector

Short-term learning for long-term impact: lessons on project design from Malawi

The complex work of sustainable development is one that benefits from applying “lessons learned” from previous experience in similar contexts. The paradox of requiring quick learning about long-term impacts can be partially resolved by co-interpretation with project partners on how a project’s impacts are expected to proceed over time, and how the project has helped and hindered sustained service delivery. In 2013, the African Development Bank (AfDB) and Engineers Without Borders Canada (EWB) used this idea to study the anticipated impacts of a past AfDB supported project in Malawi’s water, sanitation and hygiene (WASH) sector, to inform the design of a new project to be supported by AfDB. This paper describes this innovative approach to impact assessment, as well as key lessons from this study, on using decentralised structures, investing in capacity building, and building sustaining community level institutions

Community management of water points in the Democratic Republic of the Congo: identifying success factors

In the Democratic Republic of the Congo, the DRC WASH Consortium has been applying a modified Life-Cycle cost approach to improve the sustainability of rural WASH services. In particular, limited community capacity to collect funds to cover operation and maintenance costs threatens the long-term functionality of installed infrastructure: the Consortium has been supporting, since 2013, communities to develop business plans to improve the sustainability of their water points over time. This paper investigates factors characterising communities and committees that are able to reach three defined levels of financial self-sufficiency. The levels are based on the calculated costs of sustaining services over the short, medium, and long term. The factors investigated include structural factors - community size, type of water point, committee composition – and also factors associated with the operational approach of the committees – method of revenue generation, exemption of vulnerable people, and professionalisation of the committee

Codon table and the anticodon loop of tRNAs specific for Gln, Lys, and Glu which contain cmnm⁵s²U (Gln) or mnm⁵s²U (Lys and Glu) as wobble nucleoside in bacterial tRNAs.

<p>Above the third column (second codon base A) the anticodon stem and loop of the tRNAs containing the (c)mnm⁵s²U in position 34 is shown (denoted in red <b>cmnm5s2</b>). Position 36 of the anticodon is color coded with Green <b>G36</b> being position 36 for , red <b>U36</b> for , and blue <b>C36</b> for . Similar color code is denoted for the modified nucleosides present in position 37 in the corresponding tRNAs. N denotes G, A or C, respectively, for first nucleoside in the relevant codons read by these tRNAs. Note that these tRNAs are rich in U, which is a poor stacker [<a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0175092#pone.0175092.ref024" target="_blank">24</a>] making the anticodon very flexible especially if the modifications is absent and this is especially true of . In the codon table the letters outside the box, to the left, above, and to the right indicate the first, second, and third position of the codon. Circles connected by a line, or a single circle, represent one tRNA species. A filled circle indicates the capacity of that tRNA to base pair with the indicated codon, either by Watson-Crick or by wobble according to the revised wobble hypothesis [<a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0175092#pone.0175092.ref001" target="_blank">1</a>]. Red and yellow circles indicate tRNAs that are sequenced at the RNA level while green circles represent tRNAs for which only a partial tRNA sequence is available. A red or green circle indicates efficient base pairing while a yellow circle indicates a restricted wobble. An open (white) circle is a base pairing that is not according to the revised wobble hypothesis. However, data <i>in vivo</i> from mutants where only this tRNA is left to decode all codons in the codon box, suggest that the tRNA in fact is able to read that codon [<a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0175092#pone.0175092.ref009" target="_blank">9</a>, <a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0175092#pone.0175092.ref025" target="_blank">25</a>]. Data are compiled from Sprinzl data base (<a href="http://trnadb.bioinf.uni-leipzig.de/" target="_blank">http://trnadb.bioinf.uni-leipzig.de/</a> and Modomics data base (<a href="http://modomics.genesilico.pl/" target="_blank">http://modomics.genesilico.pl/</a>). Mutations in <i>mnmE</i> or <i>mnmG</i> genes result in no formation of the (c)mnm⁵- group not only of the (c)mnm⁵s²U present in tRNAs specific for Gln, Lys and Glu but also cmnm⁵Um in and mnm⁵U34 in tRNAs specific for Arg and Gly. Mutations in <i>mnmA</i> results in no thiolation of cmnm⁵s²U34. <b>a</b>) The modifications in position 34 of from <i>S</i>. <i>enterica</i> are cmnm⁵s²U (80%) and mnm⁵s²U (20%) [<a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0175092#pone.0175092.ref010" target="_blank">10</a>] and similar in <i>E</i>. <i>coli</i> [<a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0175092#pone.0175092.ref011" target="_blank">11</a>] <b>b</b>) The majority of this tRNA^Gly contains mnm⁵U34 but there is also a small amount of cmnm⁵U34 [<a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0175092#pone.0175092.ref001" target="_blank">1</a>].</p

Amino acid sequence of the frameshift product encoded from plasmids pUST290, pUST292, pUST310, and pUST311.

<p>The frameshift window, within which the frameshift must occur, is bordered by the stop codon UAA (italics and underlined) in +1 frame and the stop codon UGA UAA in the zero frame (Indicated by a * below the DNA sequence). P or F (in red) denote the last amino acid decoded in the zero frame found in the frameshift product.</p

Analysis of some typical mutations in genes inducing suppression of frameshift mutations.

<p><b>a)</b>Monitored as the ability to suppress the <i>his</i>-allele the mutant was selected to suppress. Growth of mutants on a plate lacking His following and incubation at 37°C for 4–6 days. The parental strain, which has no suppressor mutation but the indicated <i>his</i>-allele, did not grow on plates without His.</p><p><b>b)</b>The suppression was monitored as the ability to suppress the CCC-CAA-UAG sequence placed in front of the <i>lacZ</i> gene (See M–M).</p><p><b>c)</b>Plasmids (See <a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0060246#pone-0060246-g004" target="_blank">figure 4</a>) used to determine the amino acid sequence at the frameshifting site and the last amino acid in the zero frame is indicated in parenthesis.</p><p><b>d</b>)P-site according to Qian et al 1998 <a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0060246#pone.0060246-Qian1" target="_blank">[34]</a>.</p><p><b>e</b>)P-site according to <a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0060246#pone.0060246-Qian2" target="_blank">[35]</a>.</p><p><b>f</b>)According to <a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0060246#pone.0060246-Nsvall1" target="_blank">[36]</a>.</p><p><b>g)</b>According to <a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0060246#pone.0060246-Chen1" target="_blank">[69]</a>.</p><p><b>h</b>)According to <a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0060246#pone.0060246-Urbonavicius1" target="_blank">[16]</a>.</p><p><b>i</b>)P-site since these mutants also lack the s²-group of mnm⁵s²U similar to the <i>mnmA</i> mutants.</p><p><b>j)</b>P-site since <i>mnmG</i> mutants like <i>mnmE</i> mutants lack the mnm⁵-side chain of mnm5s2U.</p><p><b>k)</b>According to <a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0060246#pone.0060246-Li1" target="_blank">[71]</a>.</p><p>pUST136: contains the <i>metT</i> operon in which <i>glnU</i> is present. It also contains the gene <i>miaB</i>. See Esberg et al 1999 <a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0060246#pone.0060246-Esberg1" target="_blank">[85]</a>.</p><p>pCBS4 (<i>glnS⁺</i>) contains the <i>glnS⁺</i> wild type allele.</p

Summary of all mutants selected as suppressors to various frameshift mutations in the <i>his</i>-operon^a.

<p>a)The various <i>his</i>-alleles used were: <i>hisD3737</i> (CCC-CAA), <i>hisC10106</i> (CCC-AUG), <i>hisC10109</i> (CCC-UGG), <i>hisD10110</i> (CCC-UAU), <i>hisD10111</i> (CCC-AAG), or <i>hisD10122</i> (CCC-CAA).</p><p>b)These mutations were not sequenced but localized by transduction to a transposon closely linked to the indicated gene. The linkage between the transposon and the mutation was consistent with the mutation being in the indicated gene.</p

HPLC analysis of tRNA from <i>mnmE17<>Km</i> and <i>mnmA16<>cat</i> mutants grown in rich medium at 37°C.

<p>(<b>A</b>) Strain GT7132 (<i>mnmA</i>^<i>+</i>, <i>mnmE</i>^<i>+</i>). (<b>B</b>) Strain GT8173 (<i>mnmA16</i><>cat). (<b>C</b>) Strain GT8176 (<i>mnmE17</i><>kan). (<b>D)</b> Strain GT7132 (<i>mnmA</i>^<i>+</i>, <i>mnmE</i>^<i>+</i>). (<b>E</b>) Strain GT8173(<i>mnmA16</i><>cat). (<b>F</b>) Strain GT8176 (<i>mnmE17</i><>kan). Panel A,B and C are monitored at 254 nm and panel D,E and F are monitored at 314 nm. <i>AU</i>, absorbance units.</p

The ribosomal grip of the peptidyl-tRNA is pivotal in reading frame maintenance.

<p>The figure shows three ways (A, B and C) how certain events may induce slippage by the peptidyl-tRNA and thereby a frameshift error. It is the ternary complex (aa-tRNA*EfTu*GTP) which enters the A-site and interacts with the codon but in the figure we have symbolized it with “aa-tRNA” to save space. <b>A</b>. A defective cognate tRNA (red diamond) is slow (broken arrow) entering the A-site allowing a near-cognate aa-tRNA (blue wobble nucleoside) to decode the A-site codon. After a 3 nucleotide translocation the near-cognate peptidyl-tRNA may slip into the +1 frame. <b>B</b>. A defective cognate aa-tRNA (red diamond) decodes efficiently the codon in the A-site. After a 3 nucleotide translocation the defective cognate peptidyl-tRNA may be prone to slip into the +1 frame. <b>C</b>. The defective aa-tRNA (red diamond, yellow tRNA) is slow entering the A-site mediating a pause allowing the cognate wild type peptidyl-tRNA to slip into the +1 frame. Not depicted in the figure, alterations in the ribosomal P-site environment may also induce a frameshift error if the alteration changes the ribosomal grip of the peptidyl-tRNA. The figure is adopted from <a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0060246#pone.0060246-Nsvall1" target="_blank">[36]</a> with permission. Indeed, as shown in this paper a truncation of ribosomal protein S9, which interacts with the peptidyl-tRNA induces an error in reading frame maintenance (See Fig. 6). Moreover, the occupancy of the E-site also improves reading frame maintenance <a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0060246#pone.0060246-Jenner1" target="_blank">[80]</a>, <a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0060246#pone.0060246-Mrquez1" target="_blank">[86]</a>–<a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0060246#pone.0060246-Liao1" target="_blank">[88]</a>, perhaps by strengthening the ribosomal grip of the peptidyl-tRNA. Therefore, a defective tRNA may also increase frameshifting by altering the dissociation rate of it from the E-site.</p