Hb 201 analyser

The blood haemoglobin concentration was measured using a HemoCue Hb 201 analyser (HemoCue AB, Ängelholm, Sweden). The serum concentration of leptin was measured using a commercially available ELISA kit, following the manufacturer’s instructions (Insight Biotechnology, UK). Mice were examined for evidence of microbial infection using 2 immunocomb kits (Biogal Galed Labs, Israel), which detect antibodies to the Corona, Mouse Hepatitis, Sendai, Minute, Noro, and Parvo viruses and to M. pulmonis. These tests were scored on a 0 to 4 scale, where 0 is the absence of a response, and 1, 2, 3, and 4 are all seropositive results in increasing degrees of positivity. These microbial infections are likely to only represent a subset of all the microbial infections to which wild mice are exposed, many of which may be unknown infections.

The authors did not collect blood specimens for anaemia testing for this study. These were collected under NFHS-5 by health investigators from eligible men aged 15 to 54 with their consent. Blood samples were drawn from a drop of blood taken from a finger prick (or a heel prick for children age 6–11 months) [8 ] and collected in a microcuvette, a single-use pipette. Concentration of haemoglobin was analysed on-site with HemoCue Hb 201+ analyser. Introduced in 1990, the HemoCue Hb 201+ is a battery-operated portable device used for quantitative determination of haemoglobin level in undiluted, capillary or venous blood. It converts the haemoglobin into methemoglobin and combines it with azide to form azidemethemoglobin followed by measurement of transmittance and haemoglobin absorbance [22 (link)–24 (link)].

Daily surveillance was carried out at each health facility during regular opening hours (Monday to Friday, 7.45 am-4.06 pm). Outpatients presenting with fever or a history of fever, in the past 3 days, were referred to a study nurse who then completed an mRDT, prepared a thick and thin blood smear on a microscopy slide and measured haemoglobin (Hb) level using a Hemocue Hb 201+ analyser (HemoCue AB, Ängelholm, Sweden). Demographic details of the patient were recorded in a one-page case report form alongside clinical signs and symptoms, previous health facility attendance and drug intake, axillary temperature, weight and results of the mRDT and Hb measurement. The patient was then transferred to a member of the respective facility’s health work force for further examination, diagnosis and treatment following routine procedures. The resulting diagnosis and any prescription by the health worker were recorded on the case report form. Oral informed consent was obtained from all patients prior to participation.

In NFHS-5, iInvestigators collected blood specimens for anaemia testing from eligible men aged 15–54 years. Prior to the collection, respondents’ consent for the test was obtained. The blood samples were obtained through a finger prick method and were collected in micro cuvettes. On-site analysis of haemoglobin levels was performed using a battery-operated portable HemoCue Hb 201 + analyser. Additionally, haemoglobin levels were adjusted for altitude in enumeration areas situated at altitudes above 1,000 m to account for potential altitude-related variations [19 ].

We collected the samples from mid-April to July, during breeding seasons of 2015 and 2016. We visited research area every day to localise singing males and search for nests. We marked the nestlings individually using numbered metal rings, and measured them when they were 6–7 days old. We weighed the nestlings using an electronic scale (to the nearest 0.01 g) and measured the concentration of haemoglobin using a portable haemoglobinometer (HemoCue Hb 201+ Analyser, Sweden). We took the blood samples (10–20 μL) by puncture from the wing vein using a sterile needle and a Pasteur pipette (Arctander 1988 (link)). Blood sampling was approved by the II Local Ethics Committee in Wrocław, Poland, under number 40/2014. If defecation occurred during handling, we collected the excreta immediately in clean labelled ziplock bags. We kept the samples in the freezer, until laboratory analysis. We recorded locations of the nests by a handheld Garmin GPS device and used Quantum GIS 2.14.1. to measure the distance between the nests and the slag dump, as well as the power and heat plant. After two seasons, we collected complete data (excreta, blood and body mass) from 29 breeding nests with 2–6 nestlings each (116 nestlings in total).

A trained nurse collected blood samples through vein puncture from consented participants. Blood samples were taken into ethylenediaminetetraacetic acid (EDTA) and non-anticoagulated whole blood vacutainers (Becton Dickenson, NJ, USA). Approximately 6mL of venous blood sample was collected on each vacutainer and protected from light. Whole-blood vacutainers were maintained at 4–8°C for less than 2 hours before being transported to the temporary laboratories. Malaria was tested by rapid diagnostic test (SD Bioline, Rep. of Korea), and hemoglobin level was measured by HemoCue HB 201+ analyser (Hemo Cue, Angelholm, Sweden). Assessment of C-reactive protein (CRP), and alpha-1 acid glycoprotein (AGP) was performed with Roche Cobas Integra 400 Plus analyser (Roche Diagnostics GmbH, German). Hemoglobin levels <12.0 and <8.0 g/dL were used to characterise anaemia and severe anaemia, respectively. Serum C- reactive protein (CRP) and Alpha-1-acid glycoprotein (AGP) values of CRP > 5.0 mg/L and AGP > 1.0 g/L respectively were characterized as high inflammatory marks [26 , 27 (link)].