Growth of Gram-Negative Bacteria in Antiseptics, Disinfectants and Hand Hygiene Products in Two Tertiary Care Hospitals in West Africa-A Cross-Sectional Survey.

Antiseptics, disinfectants, and hand hygiene products can act as reservoirs of Gram-negative bacteria causing healthcare-associated infections. This problem is rarely documented in low- and middle-income countries, particularly in sub-Saharan Africa. In a cross-sectional survey, we assessed the bacterial contamination of antiseptics, disinfectants, and hand hygiene products in two university hospitals in Burkina Faso and Benin. During ward visits and staff interviews, in-use products were cultured for the presence of Gram-negative bacteria. The growth of Gram-negative bacteria was absent or rare in alcohol-based products, povidone iodine, and Dakin solution. Contamination was highest (73.9% (51/69)) for liquid soap products (versus antiseptic/disinfectants (4.5%, 7/157) (p < 0.0001)), mostly used in high-risk areas and associated with high total bacterial counts (>10,000 colony-forming units/mL). Contaminating flora (105 isolates) included Enterobacterales and the Vibrio non-cholerae/Aeromonas group (17.1%) and non-fermentative Gram-negative rods (82.8%). Multidrug resistance was present among 9/16 Enterobacterales (Klebsiella and Enterobacter spp.) and 3/12 Acinetobacter spp., including carbapenem resistance (Acinetobacter baumannii: NDM, Pseudomonas stutzeri: VIM). The risk factors for contamination included the type of product (cleaning grade and in-house prepared liquid soap), use of recycled disposable containers and soft drink bottles, absence of labeling, topping-up of containers, dilution with tap water (pharmacy and ward), and poor-quality management (procurement, stock management, expiry dates, and period after opening).

Salmonella Typhi whole genome sequencing in Rwanda shows a diverse historical population with recent introduction of haplotype H58.

Salmonella enterica serovar Typhi (S. Typhi) is the cause of typhoid fever, presenting high rates of morbidity and mortality in low- and middle-income countries. The H58 haplotype shows high levels of antimicrobial resistance (AMR) and is the dominant S. Typhi haplotype in endemic areas of Asia and East sub-Saharan Africa. The situation in Rwanda is currently unknown and therefore to reveal the genetic diversity and AMR of S. Typhi in Rwanda, 25 historical (1984-1985) and 26 recent (2010-2018) isolates from Rwanda were analysed using whole genome sequencing (WGS). WGS was locally implemented using Illumina MiniSeq and web-based analysis tools, thereafter complemented with bioinformatic approaches for more in-depth analyses. Whereas historical S. Typhi isolates were found to be fully susceptible to antimicrobials and show a diversity of genotypes, i.e 2.2.2, 2.5, 3.3.1 and 4.1; the recent isolates showed high AMR rates and were predominantly associated with genotype 4.3.1.2 (H58, 22/26; 84,6%), possibly resulting from a single introduction in Rwanda from South Asia before 2010. We identified practical challenges for the use of WGS in endemic regions, including a high cost for shipment of molecular reagents and lack of high-end computational infrastructure for the analyses, but also identified WGS to be feasible in the studied setting and giving opportunity for synergy with other programs.

Urban rats as carriers of invasive Salmonella Typhimurium sequence type 313, Kisangani, Democratic Republic of Congo.

BACKGROUND: Invasive non-typhoidal Salmonella (iNTS-mainly serotypes Enteritidis and Typhimurium) are major causes of bloodstream infections in children in sub-Saharan Africa, but their reservoir remains unknown. We assessed iNTS carriage in rats in an urban setting endemic for iNTS carriage and compared genetic profiles of iNTS from rats with those isolated from humans. METHODOLOGY/PRINCIPAL FINDINGS: From April 2016 to December 2018, rats were trapped in five marketplaces and a slaughterhouse in Kisangani, Democratic Republic of the Congo. After euthanasia, blood, liver, spleen, and rectal content were cultured for Salmonella. Genetic relatedness between iNTS from rats and humans-obtained from blood cultures at Kisangani University Hospital-was assessed with multilocus variable-number tandem repeat (VNTR) analysis (MLVA), multilocus sequence typing (MLST) and core-genome MLST (cgMLST). 1650 live-capture traps yielded 566 (34.3%) rats (95.6% Rattus norvegicus, 4.4% Rattus rattus); 46 (8.1%) of them carried Salmonella, of which 13 had more than one serotype. The most common serotypes were II.42:r:- (n = 18 rats), Kapemba (n = 12), Weltevreden and Typhimurium (n = 10, each), and Dublin (n = 8). Salmonella Typhimurium belonged to MLST ST19 (n = 7 rats) and the invasive ST313 (n = 3, isolated from deep organs but not from rectal content). Sixteen human S. Typhimurium isolates (all ST313) were available for comparison: MLVA and cgMLST revealed two distinct rat-human clusters involving both six human isolates, respectively, i.e. in total 12/16 human ST313 isolates. All ST313 Typhimurium isolates from rats and humans clustered with the ST313 Lineage 2 isolates and most were multidrug resistant; the remaining isolates from rats including S. Typhimurium ST19 were pan-susceptible. CONCLUSION: The present study provides evidence of urban rats as potential reservoirs of S. Typhimurium ST313 in an iNTS endemic area in sub-Saharan Africa.

Use of WATCH antibiotics prior to presentation to the hospital in rural Burkina Faso.

BACKGROUND: In low- and middle-income countries, the prevalence of antimicrobial resistance (AMR) is increasing. To control AMR, WHO recommends monitoring antibiotic use, in particular Watch antibiotics. These are critically important antibiotics, with restricted use because at risk of becoming ineffective due to increasing AMR. We investigated pre-hospital antibiotic use in rural Burkina Faso. METHODS: During 2016-2017, we collected data from patients aged > 3 months presenting with severe acute fever to the rural hospital of Nanoro Health District, Burkina Faso, including antibiotic use in the two weeks prior to consultation or hospitalization. We analysed reported antibiotic use by applying the WHO Access, Watch, Reserve classification. RESULTS: Of 920 febrile participants (63.0% ≤ 14 years), pre-hospital antibiotic use was reported by 363 (39.5%). Among these 363, microbiological diagnoses were available for 275 (75.8%) patients, of whom 162 (58.9%) were non-bacterial infections. Use of more than one antibiotic was reported by 58/363 (16.0%) participants. Of 491 self-referred patients who did not previously visit a primary health care center, 131 (26.7%) reported antibiotic use. Of 424 antibiotics reported, 265 (62.5%) were Access and 159 (37.5%) Watch antibiotics. Watch antibiotic use was more frequent among patients > 14 year olds (51.1%) compared to those 0-14 year old (30.7%, p < 0.001) and among referrals from the primary health care centers (42.2%) compared to self-referred patients (28.1%, p = 0.004). Most frequently reported Watch antibiotics were ceftriaxone (114, 71.7%) and ciprofloxacin (32, 20.1%). CONCLUSION: The reported frequent use of Watch group antibiotics among febrile patients prior to presentation to the hospital in rural Burkina Faso highlights the need to develop targeted interventions to improve antibiotic use in community settings as part of strengthening antibiotic stewardship in low- and middle-income countries. This should include facilitating referral, access to qualified prescribers and diagnostic tools in rural primary health care centers. Trial registration ClinicalTrials.gov identifier: NCT02669823. Registration date was February 1, 2016.

Aetiologies of acute undifferentiated febrile illness at the emergency ward of the University of Gondar Hospital, Ethiopia.

OBJECTIVE: Causes of acute febrile illness (AFI) often remain undetermined in developing countries, due to overlap of symptoms and limited available diagnostics. We aimed to assess the aetiology of AFI in adults in a referral hospital in northwest Ethiopia. METHODS: While all participants were tested for malaria by rapid diagnostic test (RDT), microscopy was only done on physician's request. Dengue virus (DENV) infections were detected using an RDT and ELISAs and dengue, yellow fever and chikungunya cases were identified by PCR. Bacterial aetiologies were investigated using blood culture and PCR. RESULTS: The aetiology of acute infection was identified for 20.5% of 200 patients enrolled. Eleven percent tested positive for Plasmodium, while microscopy was only requested for half of the identified malaria cases. For 4.0% of the Plasmodium-infected patients, an acute or past DENV (co-)infection was detected. We found 7.5% acute and 13.0% past DENV - all serotype 3 - infections. Bacterial infections were observed in 4.5% of the patients. CONCLUSION: Malaria is still a considerable aetiology of AFI and dengue is underrecognised. There are areas where both diseases occur concomitantly, and the DENV-3 serotype presumably spreads from Sudan to northern Ethiopia. As only 20.5% of the aetiologies were identified, a broader testing platform is required.

Underutilization and Quality Gaps in Blood Culture Processing in Public Hospitals of Peru.

Correct processing of blood cultures may impact individual patient management, antibiotic stewardship, and scaling up of antimicrobial resistance surveillance. To assess the quality of blood culture processing, we conducted four assessments at 16 public hospitals across different regions of Peru. We assessed the following standardized quality indicators: 1) positivity and contamination rates, 2) compliance with recommended number of bottles/sets and volume of blood sampled, 3) blood culture utilization, and 4) possible barriers for compliance with recommendations. Suboptimal performance was found, with a median contamination rate of 4.2% (range 0-15.1%), with only one third of the participating hospitals meeting the target value of < 3%; and a median positivity rate of 4.9% (range 1-8.1%), with only 6 out of the 15 surveilled hospitals meeting the target of 6-12%. None of the assessed hospitals met both targets. The median frequency of solitary blood cultures was 71.9% and only 8.9% (N = 59) of the surveyed adult bottles met the target blood volume of 8 - 12 mL, whereas 90.5% (N = 602) were underfilled. A high frequency of missed opportunities for ordering blood cultures was found (69.9%, 221/316) among patients with clinical indications for blood culture sampling. This multicenter study demonstrates important shortcomings in the quality of blood culture processing in public hospitals of Peru. It provides a national benchmark of blood culture utilization and quality indicators that can be used to monitor future quality improvement studies and diagnostic stewardship policies.

Nonautomated Blood Cultures in a Low-Resource Setting: Optimizing the Timing of Blind Subculture.

Laboratory procedures for blood cultures in a hospital in Phnom Penh were adapted to optimize detection of Burkholderia pseudomallei, an important pathogen in this setting. The effects of these changes are analyzed in this study. Blood cultures consisted of two BacT/ALERT bottles (bioMérieux, Marcy-l'Etoile, France). Growth was detected visually by daily inspection of the bottles. In 2016, the aerobic-anaerobic pair (FA/FN FAN) was substituted by an aerobic pair of BacT/ALERT FA Plus bottles. Blind subculture (BS) (subculture in the absence of visual growth) was advanced from day 3 to day 2 of incubation in July 2016. In July 2018, it was further advanced to day 1 of incubation. From July 2016 to October 2019, 9,760 blood cultures were sampled. The proportion of cultures showing pathogen growth decreased from 9.6% to 6.8% after the implementation of the laboratory changes (P < 0.001). Advancing the BS from day 3 to day 2 led to an increased proportion of pathogens detected by day 3 (92.8% versus 82.3%; P < 0.001); for B. pseudomallei, this increase was even more remarkable (92.0% versus 18.2%). Blind subculture on day 1 similarly increased the proportion of pathogens detected by day 2 (82.9% versus 69.0% overall, 66.7% versus 10.0% for B. pseudomallei; both P < 0.001). However, after implementation of day 1 subculture, a decrease in recovery of B. pseudomallei was observed (12.4% of all pathogens versus 4.3%; P < 0.001). In conclusion, earlier subculture significantly shortens time to detection and time to actionable results. Some organisms may be missed by performing an early subculture, especially those that grow more slowly.

Slow growth of Burkholderia pseudomallei compared to other pathogens in an adapted blood culture system in Phnom Penh, Cambodia.

PURPOSE: Burkholderia pseudomallei is a key pathogen causing bloodstream infections at Sihanouk Hospital Center of Hope, Phnom Penh, Cambodia. Here, visual instead of automated detection of growth of commercial blood culture bottles is done. The present study assessed the performance of this system. METHODOLOGY: Blood culture sets, consisting of paired adult aerobic and anaerobic bottles (bioMérieux, FA FAN 259791 and FN FAN 252793) were incubated in a standard incubator for 7 days after reception. Each day, the bottle growth indicator was visually inspected for colour change indicating growth. Blind subculture was performed from the aerobic bottle at day 3. RESULTS: From 2010 to 2015, 11  671 sets representing 10  389 suspected bloodstream infection episodes were documented. In 1058 (10.2  %) episodes, pathogens grew; they comprised Escherichia coli (31.7 %), Salmonella Paratyphi A (13.9 %), B. pseudomallei (8.5 %), Staphylococcus aureus (7.8 %) and Klebsiella pneumoniae (7.0 %). Blind subculture yielded 72 (4.1  %) pathogens, mostly (55/72, 76.4 %) B. pseudomallei. Cumulative proportions of growth at day 2 were as follows: E. coli: 85.0 %, Salmonella Paratyphi A: 85.0 %, K. pneumoniae: 76.3  % and S. aureus: 52.2  %; for B. pseudomallei, this was only 4.0  %, which increased to 70.1  % (70/99) at day 4 mainly by detection on blind subculture (55/99). Compared to the anaerobic bottles, aerobic bottles had a higher yield and a shorter time-to-detection, particularly for B. pseudomallei. CONCLUSIONS: Visual inspection for growth of commercial blood culture bottles in a low-resource setting provided satisfactory yield and time-to-detection. However, B. pseudomallei grew slowly and was mainly detected by blind subculture. The aerobic bottle outperformed the anaerobic bottle.

Diagnostic accuracy of the InBiOS AMD rapid diagnostic test for the detection of Burkholderia pseudomallei antigen in grown blood culture broth.

To assess the diagnostic and operational performance of the InBiOS AMD rapid diagnostic test (RDT) (Seattle, USA) for the detection of B. pseudomallei in grown blood culture broth. The InBiOS RDT is a lateral flow immunoassay in a strip format detecting B. pseudomallei capsular polysaccharide in culture fluids, marketed for research only. Broth of blood culture bottles (BacT/Alert, bioMérieux, Marcy L'Etoile, France) sampled in adult patients at the Sihanouk Hospital Center of HOPE, Phnom Penh, Cambodia, during 2010-2017 and stored at - 80 °C was tested. They included samples grown with B. pseudomallei (n = 114), samples with no growth (n = 12), and samples with growth of other pathogens (n = 139, among which Burkholderia cepacia (n = 5)). Diagnostic sensitivity and specificity were 96.5% [95% confidence interval (CI): 91.3-98.6%] and 100% [CI: 97.5-100%] respectively. Background clearance and line intensities were good and very good. The RDT's test strip, not housed in a cassette, caused difficulties in manipulation and biosafety. The centrifugation step prescribed by the procedure challenged biosafety, but processing of 19 B. pseudomallei samples without centrifugation showed similar results for line intensity and background clearance, compared to centrifugation. The InBiOS RDT showed excellent accuracy for detection of B. pseudomallei in grown blood culture broth. Provided operational adaptations such as cassette housing, it has the potential to reduce time to diagnosis of melioidosis.

Diagnostic accuracy of antigen-based immunochromatographic rapid diagnostic tests for the detection of Salmonella in blood culture broth.

BACKGROUND: In low resource settings, Salmonella serovars frequently cause bloodstream infections. This study investigated the diagnostic performance of immunochromatographic rapid diagnostic tests (RDTs), which detect Salmonella antigens, when applied to stored grown blood culture broth. MATERIAL/METHODS: The SD Bioline One Step Salmonella Typhi Ag Rapid Detection Kit (Standard Diagnostics, Republic of Korea), marketed for the detection of Salmonella enterica serovar Typhi (Salmonella Typhi) in stool and the Salmonella Ag Rapid Test (Creative Diagnostics, USA), marketed for the detection of all Salmonella serotypes in stool, were selected for evaluation based on a pre-test evaluation of six RDT products. The limits of detection (LOD) for culture suspensions were established and the selected RDT products were assessed on 19 freshly grown spiked blood culture broth samples and 413 stored clinical blood culture broth samples, collected in Cambodia and the Democratic Republic of the Congo. RESULTS: The LOD of both products was established as 107-108 CFU/ml. When applied to clinical blood culture broth samples, the diagnostic sensitivity and specificity of the SD Bioline RDT were respectively 100% and 79.7% for the detection of Salmonella Typhi; 94.4% (65/69) of false-positive results were caused by Salmonella Enteritidis. When considering the combined detection of Salmonella Typhi and Enteritidis (both group D Salmonella), sensitivity and specificity were 97.9% and 98.5% respectively. For Creative Diagnostics, diagnostic sensitivity was 78.3% and specificity 91.0% for all Salmonella serotypes combined; 88.3% (53/60) of false negative results were caused by Salmonella Paratyphi A. CONCLUSIONS: When applied to grown blood culture broths, the SD Bioline RDT had a good sensitivity and specificity for the detection of Salmonella Typhi and Salmonella Enteritidis. The Creative Diagnostics product had a moderate sensitivity and acceptable specificity for the detection of all Salmonella serovars combined and needs further optimization. A RDT that reliably detects Salmonella Paratyphi A is needed.