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Co-infection best predicts respiratory viral infection in a wild host

Citation

Glidden, Caroline et al. (2020), Co-infection best predicts respiratory viral infection in a wild host, Dryad, Dataset, https://doi.org/10.5061/dryad.hmgqnk9fj

Abstract

1) The dynamics of directly transmitted pathogens in natural populations are likely to result from the combined effects of host traits, pathogen biology and interactions among pathogens within a host. Discovering how these factors work in concert to shape variation in pathogen dynamics in natural host – multi‐pathogen systems is fundamental to understanding population health.

2) Here, we describe temporal variation in incidence and then elucidate the effect of hosts trait, season, and pathogen co‐occurrence on host infection risk using one of the most comprehensive studies of co‐infection in a wild population: a suite of seven directly‐transmitted, viral and bacterial, respiratory infections from a four‐year study of 200 free‐ranging African buffalo (Syncerus caffer).

3) Incidence of upper respiratory infections was common throughout the study – five out of the seven pathogens appeared to be consistently circulating throughout our study population. One pathogen exhibited clear outbreak dynamics in our final study year and another was rarely detected.

4) Co‐infection was also common in this system. The strongest indicator of pathogen occurrence for respiratory viruses was, in fact, the presence of other viral respiratory infections. Host traits had minimal effects on odds of pathogen occurrence but did modify pathogen‐pathogen associations. In contrast, only season predicted bacterial pathogen occurrence.

5) Though a combination of environmental, behavioral, and physiological factors work together to shape disease dynamics, we found pathogen associations best determined infection risk. Our study demonstrates that, in absence of very fine‐scale data, the intricate changes among these factors are best represented by co‐infection.

Methods

Materials and methods (from JAE https://doi.org/10.1111/1365-2656.13391)

2.1 Study area

Kruger National Park (KNP) is located in the north-eastern corner of South Africa between 22.5 and 25.5°S, and 31.0 and 31.6°E (SI Figure 1). The area of the KNP is 19,485 km2, but since 2002, the area available to wildlife has effectively doubled due to the removal of fences between private game reserves in the west and Mozambique in the east. The population of African buffalo in the park is about 37,000 animals (SANPARKS 2010-2011). Our four-year project was restricted to buffalo in the southern KNP and took place between June 2008 and June 2012.

On average, 84% of KNP’s total rainfall is concentrated between November to April (Zambatis 2003) with approximately 600 mm of rainfall per year in the southern KNP (Venter & Gertenbach 1986). The dry season typically occurs May – October.  Rather than using calendar year in our analyses, we used rainfall year, hereafter referred to as “year,” with year commencing in November.

 

2.2 Sampling regime

Female African buffalo between 2-5 years old were captured as part of a study on parasite interactions in free-ranging buffalo (Ezenwa & Jolles 2015). The first 100 buffalo were captured from the Lower Sabie herd between 23 June and 5 July, 2008 (SI figure 1). The second 100 buffalo were captured from the Crocodile Bridge herd between 1 and 8 October, 2008 (SI Figure 1). Buffalo were re-captured approximately every 6 months after this initial capture, through June 2012. Any buffalo that died or emigrated from the study area during the study period was replaced with an animal of similar age so that a near-constant sample size of 200 was maintained at each capture (additional detail in Spaan et al., 2019).

 

2.3 Sample and data collection

Buffalo were located and identified via radio-collars (7 GPS collars to locate herds, 193 VHF collars to identify individuals). At capture, buffalo were chemically immobilized with etorphine hydrochloride (M99) and ketamine by darting from a truck or helicopter. Following sample and data collection, immobilization was reversed using diprenorphine (M5050). All immobilizations were performed by South African National Park’s (SANParks) veterinarians and registered project staff, and all procedures were approved by Oregon State University, University of Georgia and SANPark’s Institutional Animal Care and Use Committees.

While buffalo were immobilized, blood and host-trait data were collected from each animal. Blood samples for serological assays were collected via jugular venipuncture in sterile tubes containing no anticoagulant. Blood was placed on ice and stored in a cooler box within 5 minutes for transportation back to the laboratory. At the laboratory, serum was collected after centrifugation for 20 minutes at 2,000 g and stored at -20°C until analysis. Host-trait measures included age, body condition, horn width, pregnancy status, lactation status and calf-at-heel status using previously published methods (Table 2).

Half of the studied buffalo in each herd were administered an oral anthelmintic treatment in the form of a Panacur® slow-release bolus at every capture. The bolus contains the active ingredient fenbendazole. Nematode egg shedding is effectively eliminated in buffalo for ~160 days after a single administration (Ezenwa et al., 2010) and alters African buffalo response to infection by microparasites (Ezenwa & Jolles 2015). The other half of each herd did not receive Panacur® at any capture.

 

2.4 Serology

2.4.1 Serological assays

Buffalo sero-status for each of the respiratory pathogens were determined using commercially available assays after each capture, as previously described (Glidden et al., 2018). Briefly, monoclonal antibodies specific to the F protein of Bovine Respiratory Syncytial Virus (BRSV) and the NS3 protein Bovine Viral Diarrheal Virus (BVDV) were detected in serum using separate competitive ELISA kits (Bio-X Diagnostics, Belgium) while Bovine herpesvirus-1 (BHV), Pi-3 (Bovine Parainfluenza-3), AD-3 (Bovine Adenovirus-3), MB (Mycoplasma bovis), and MH (Mannheimia haemolytica) serostatuses were assessed using direct ELISA test kits (Bio-X Diagnostics, Belgium). For the first set of tests, MB and MH were not included on the assay. Buffalo were tested for bTB using the BOVIGAM ELISA kit (Prionics, Switzerland) which is a standard whole blood interferon-gamma (IFNγ) assay (Wood & Jones 2001; Schiller et al., 2009). This kit in particular has been previously optimized for use in African buffalo (Michel et al., 2011). All serum was stored and analyzed in the same laboratory (KNP Veterinary Wildlife Services).

 

2.4.2  Classifying occurrence

All animals were recruited as adults, so it was impossible to determine whether the first detected increases in antibody titers were due to a primary exposure, re-exposure or recrudescence (Combink et al., 2020). For this reason, we define pathogen “occurrence” to include all three possibilities. We expect occurrence, representing all three possibilities, to represent initiation of active, transmissible infections. Identical to Glidden et al. (2018), BRSV and BVDV were tested using a competitive ELISA which give scores of 0-100% positive. Samples were deemed positive if ELISA scores were > 50%, per manufacturer instructions.  If the animal was tested 6 or more times with only one, weakly positive (<65%) result, we assumed the test result was a false positive. Occurrence of BRSV or BVDV was counted if test results went from negative (<50%) to positive (>50%) or if positive animals had a 15+% increase in their competitive ELISA score from one capture to the next (Glidden et al., 2018). We observed low occurrence of BVDV and thus excluded it from further analyses (Fig 1). For BHV, Pi-3, AD-3, MH and MB, ELISAs were scored on a 0-5 scale and occurrence was counted if the ELISA score increased by 2 or more points between two captures, per manufacturer instructions. ELISA results were previously shown to correlate with other markers of inflammation and infection (Glidden et al., 2018).

To determine bTB conversion we considered an animal’s full IFNγ bTB data set (all sampling points): an animal was considered to have become infected with bTB (bTB+) only if we observed at least two consecutive negative tests followed by at least two consecutive positive tests was observed (Ezenwa & Jolles 2015). In total, 80 animals converted to bTB+ throughout the study.

 

Table 2. Host traits and environmental variables included in our CRF analysis.

 

Variable

Variable

type

Collection information,

data transformation, and units

Season

Categorical

Pathogen exposure regimes, pathogen viability, host behavior and host immunocompetence may fluctuate with season thus we included season at sampling to detect seasonality of pathogen occurrence and hypothesize about seasonally variables not explicitly defined within our model. Wet = Nov –  Apr;  Dry = May – Oct.

Age

Continuous

Approximate age of each animal, in months, based on teeth regressions as per Jolles, Cooper and Levin (2005).

Capture herd

Binomial

This variable refers to the herd in which the buffalo was found during the given capture, Crocodile Bridge or Lower Sabie.

Condition

Categorical

Visualization and palpation of the ribs, spine, hips and the base of the tail was scored on a scale of 1 (very poor) to 5 (excellent); overall body condition score was calculated as the average of these four scores. Condition at the beginning of the interval, i.e. at the previous capture, was used in analyses (Ezenwa, Jolles & O’Brien 2009).

Age-horn residual

Continuous

The regression residuals of age at first capture on horn width (cm) was collected at the previous capture. In female buffalo, the variable is a marker of GI parasite infection with higher residuals indicating lower parasite richness as well as lower coccidia occurrence and intensity (Ezenwa & Jolles 2008).

Pregnancy status

Binomial

Pregnancy status is based on palpation by a veterinarian via rectal palpation

Lactation status

Binomial

Lactation status was assessed by manual milking of all four teats (Jolles 2007)

Calf at heel status

Binomial

This variable indicates whether there was a calf at the mother’s side during visual surveys when animals were being picked out for darting

 bTB convert

Binomial

bTB is typically a chronic, subclinical disease of the lung and upper respiratory tract in African buffalo. bTb interactions with host traits and other pathogens have been well characterized in African buffalo. Due to the chronicity of infection, we included if the animal converted to bTb as a host trait.

Anthelmintic bolus treatment

Binomial

As part of another study (Ezenwa & Jolles 2015), half of the buffalo in each herd were administered a slow-release, oral anti-helminthic treatment (Fenbendazole aka Panacur) at every capture.

 

Usage Notes

Please see ReadMe file for full description of data. 

Funding

National Institute of Mental Health, Award: T32 OD10999

University of Pretoria

Achievement Rewards for College Scientists Foundation