Although mites of the Orthohalarachne genus are common parasites of otariids, their role as agents of disease and in causing population-level mortality is unknown. In the austral summer of 2016, there was an increase in mortality among South American fur seal (Arctocephalus australis) pups at Guafo Island, Northern Chilean Patagonia. Pups found dead or terminally ill had moderate to marked, multifocal, mucopurulent bronchopneumonia associated with large numbers of respiratory mites (Orthohalarachne diminuata) and rare Gram-positive cocci. In lung areas less affected by bronchopneumonia, acute interstitial pneumonia with marked congestion and scant hemorrhage was evident. Bacteria from pups dying of bronchopneumonia were isolated and identified as Streptococcus marimammalium and Streptococcus canis. Respiratory mites obstructed airflow, disrupted airway epithelial lining, and likely facilitated the proliferation of pathogenic β-hemolytic streptococci, leading to severe bronchopneumonia and death of fur seal pups. An abrupt increase in sea surface temperature in Guafo Island corresponded to the timing of the bronchopneumonia outbreak. The potential role of environmental factors in the fur seal pup mortality warrants further study.

Mites of the Halarachnidae family (Acarina: Gamasidae) infect the respiratory system of several mammalian groups including most pinniped species (Kim et al. 1980; Alonso-Farré et al. 2012). Although these mites can reach 100% prevalence in otariids (Kim et al. 1980), their role as agents of disease and mortality of marine mammals is not well established. Species of β-hemolytic streptococci can cause pneumonia, septicemia, endocarditis, and abscesses in marine mammals (Thornton et al. 1998).

The South American fur seal (SAFS; Arctocephalus australis) is a widespread otariid species in South America; however, Chilean populations have experienced a severe decline over the last 30 yr (Seguel et al. 2013). The reasons for this decline are still unknown but major causes of mortality include entanglement, starvation, and hookworm disease (Seguel et al. 2013, 2017). As part of a long-standing health monitoring program, SAFS pup mortality has been monitored at Guafo Island, Northern Chilean Patagonia (43°35′35″S, 74°42′49″W) since 2012 (Seguel et al. 2013). In February 2016, an increase in mortality was evident, including two terminally ill pups that had loss of consciousness and were in a semiconvulsive state.

Blood sample collection and complete blood cell counts were performed in these animals as previously described (Seguel et al. 2016). Moribund pups were humanely euthanized by sedation with xylazine (Centrovet®, Santiago, Chile, 1.5 mg/kg, intramuscular) followed by euthanasia solution (T-61®, Merck, Whitehouse Station, New Jersey, USA). Complete necropsies were performed on these pups as well as on seven fresh pup carcasses collected during the last 2 wk of February 2016. For comparison, necropsies were performed on 13 fresh carcasses collected during 2017, when no unusual mortalities had occurred. Mortality rates between years were compared through a chi-square test with significance at α=0.05 using R software (R Development Core Team 2017). Tissue collection and processing for histopathologic examination were performed as previously described (Seguel et al. 2011, 2017). Sterile culture swabs from lung, trachea, bronchi, brain, liver, spleen, mesenteric lymph node, and intestine were collected and cultured individually using standardized methods (Seguel et al. 2017). To correctly identify isolated bacteria by species, DNA was extracted from pure colonies using a commercial kit (EZNA®, Omega Bio-Tek, Norcross, Georgia, USA) and was subsequently used for amplification and sequencing of about 1,500 base pairs of the bacterial 16S ribosomal RNA gene (Montagner et al. 2010). Mites were retrieved from the nasal cavity, trachea, and bronchi by collecting the mucoid exudate on mucosal surfaces and washing with saline on a 400-μm mesh. Mites were fixed in 70% ethanol, cleared with Nesbitt solution, mounted in a Berlese's medium, and examined under optical microscopy for detailed morphologic identification following standard keys (Newell 1947; Gastal et al. 2016).

The pup mortality rate was significantly higher in 2016 (12.5%, 125/998) compared to 2017 (7.8%, 96/1230, χ2=11.21, df=1, P<0.001). Complete blood cell counts and serum chemistries of the two sick pups found alive revealed moderate leukopenia with marked lymphopenia and left-shift, mild hypoalbulinemia, moderate hyperglobulinemia, and moderately elevated alkaline phosphatase (Table 1) according to normal values described for SAFS pups (Seguel et al. 2016). All pups necropsied in 2016 were in good nutritional condition. The most-significant gross findings we found included diffusely dark-red lungs and a large amount of mucopurulent material in the trachea and bronchi admixed with numerous, 5-mm long white mites identified as Orthohalarachne diminuata (Fig. 1A, B). Multifocally, between 40% and 50% of the lung was consolidated. Histologically, 500-μm long mites with birefringent exoskeleton, striated muscle, coelom, and reproductive tract–obstructed medium-sized bronchi (Fig. 1C). In these areas, the respiratory epithelium was nonciliated, flattened, or eroded. Adjacent airways were filled with PAS-Alcian blue-positive mucus admixed with numerous degenerate neutrophils, cellular debris, and rare Gram-positive cocci colonies (Fig. 1D). In adjacent areas, approximately 20% of alveoli contained rare, degenerate leukocytes, scant cellular debris, and rare erythrocytes. Alveolar capillaries were engorged and small numbers of macrophages, lymphocytes, and neutrophils slightly expanded alveolar septae (Fig. 1E). In the brain, moderate edema expanded the meninges and brain stem Virchow-Robin spaces (Fig. 1F).

Table 1

Relevant complete blood cell count and serum chemistry findings in two South American fur seal (Arctocephalus australis) pups at Guafo Island, Northern Chilean Patagonia infected with Orthohalarachne diminuata and Streptococcus marimammalium in 2016.

Relevant complete blood cell count and serum chemistry findings in two South American fur seal (Arctocephalus australis) pups at Guafo Island, Northern Chilean Patagonia infected with Orthohalarachne diminuata and Streptococcus marimammalium in 2016.
Relevant complete blood cell count and serum chemistry findings in two South American fur seal (Arctocephalus australis) pups at Guafo Island, Northern Chilean Patagonia infected with Orthohalarachne diminuata and Streptococcus marimammalium in 2016.
Figure 1

Respiratory mite (Orthohalarachne diminuata) and β-hemolytic streptococci bronchopneumonia in South American fur seal (Arctocephalus australis) pups at Guafo Island, Northern Chilean Patagonia. (A) A large amount of mucopurulent material occupies the tracheal lumen. (B) Numerous mites attached to the mucosa of a main bronchus. Inset: Cleared adult female specimen of O. diminuata. (C) An adult mite obstructs the lumen and compresses the epithelium of a bronchiolus. Moderate amounts of degenerate leukocytes and mucus surround the mite (asterisk). H&E. Scale bar=100 μm. (D) Large amount of mucus (asterisk) and degenerate leukocytes obstruct a major bronchus. Alcian blue-PAS. Scale bar=100 μm. Upper inset: Small colonies of cocci admixed with mucus and cellular debris. H&E. Scale bar=10 μm. Lower inset: Cocci are Gram-positive. Brown and Brenn Gram stain. Scale bar=10 μm. (E) Small numbers of macrophages and lymphocytes slightly expand alveolar septae. Alveolar capillaries are engorged. H&E. Scale bar=50 μm. (F) Moderate edema expands the cerebral meninges. H&E. Scale bar=100 μm.

Figure 1

Respiratory mite (Orthohalarachne diminuata) and β-hemolytic streptococci bronchopneumonia in South American fur seal (Arctocephalus australis) pups at Guafo Island, Northern Chilean Patagonia. (A) A large amount of mucopurulent material occupies the tracheal lumen. (B) Numerous mites attached to the mucosa of a main bronchus. Inset: Cleared adult female specimen of O. diminuata. (C) An adult mite obstructs the lumen and compresses the epithelium of a bronchiolus. Moderate amounts of degenerate leukocytes and mucus surround the mite (asterisk). H&E. Scale bar=100 μm. (D) Large amount of mucus (asterisk) and degenerate leukocytes obstruct a major bronchus. Alcian blue-PAS. Scale bar=100 μm. Upper inset: Small colonies of cocci admixed with mucus and cellular debris. H&E. Scale bar=10 μm. Lower inset: Cocci are Gram-positive. Brown and Brenn Gram stain. Scale bar=10 μm. (E) Small numbers of macrophages and lymphocytes slightly expand alveolar septae. Alveolar capillaries are engorged. H&E. Scale bar=50 μm. (F) Moderate edema expands the cerebral meninges. H&E. Scale bar=100 μm.

Close modal

In contrast, pups that died during 2017 had typical gross and histologic findings of trauma (n=5), drowning (n=2), emaciation (n=1), and hookworm enteritis and bacteremia (n=5; Seguel et al. 2011, 2017). Tracheal and lung swabs from five pups found during the bronchopneumonia outbreak in 2016, including the two euthanized pups, yielded moderate growth of Gram-positive, β-hemolytic streptococci. The bacterium was identified as Streptococcus marimammalium in four individuals whereas in one individual the bacterium was identified as Streptococcus canis. Tissue samples from the other three pups yielded no bacterial growth whereas a lung swab sample from one pup yielded mixed growths of Escherichia coli, Proteus sp., and Enterococcus faecalis, likely from field contamination. Swabs collected from the lung, spleen, liver, and lung of pups found dead during 2017 yielded either pure cultures of E. coli (n=3; animals died of hookworm enteritis and bacteremia) or mixed growth of Staphylococcus equorum and Staphylococcus intermedius (n=2; animals died of trauma), or no bacterial growth (n=8).

The assessment of moribund animals, and the gross and histopathology findings on fresh carcasses collected during an SAFS mortality event at the end of the 2016 breeding season, indicated that pups most likely died because of bronchopneumonia and sepsis associated with S. canis or S. marimammalium infection and a heavy burden of O. diminuata. The heavy respiratory mite infection could have predisposed the pups to the colonization and proliferation of streptococci, which may have increased the severity of the bronchopneumonia and led to respiratory distress and potentially to sepsis and death. In previous reproductive seasons, 30% to 60% of SAFS pups necropsied at the end of the reproductive season were infected with O. diminuata (Seguel et al. 2011); in 2016, all necropsied pups were infected. Additionally, the infection burden in 2016 was apparently higher when compared to previous years and to 2017 based on semiquantitative scoring during necropsies (Seguel et al. 2011). Transmission of respiratory mites probably occurs through direct contact with infected adult animals because mite nymphs and larvae inhabit the host nasal cavity while adult mites infect the lower respiratory tract (Kim et al. 1980). The substantial damage to the respiratory mucosa and obstruction of airways of pups examined in 2016 could be attributable to a density-dependent effect of the mites. In this scenario, it is likely that the mucociliary system was impaired, favoring proliferation of bacteria in small airways. Additionally, the movement of nymphs and larvae from the nasal cavity to the lung to complete their life cycle may have transferred bacteria from the upper to the lower respiratory tract.

We know that β-hemolytic streptococci species cause pneumonia, septicemia, endocarditis, and abscesses in marine mammals. For example, Streptococcus phocae is one of the most-commonly isolated species (Thornton et al. 1998). Although S. marimammalium has been found incidentally in the respiratory tract of grey seals (Halichoerus grypus) and common seals (Phoca vitulina), little is known of its pathogenic potential (Lawson et al. 2005). In contrast, S. canis is a common cause of pneumonia and septicemia in domestic dogs (Canis lupus familiaris), especially in animals with comorbidities (Fulde and Valentin-Weigand 2012). Superantigens produced by group-A S. canis bind to major histocompatibility complex-II molecules, thereby eliciting a cytokine storm that may lead to hypotension, coagulopathy, shock, and multiple organ failure (Fulde and Valentin-Weigand 2012). Therefore, it is possible that the observed pulmonary congestion, hemorrhage, and the meningeal and brainstem edema are consequences of vascular leakage elicited by streptococcal toxins and inflammatory cytokines. On the other hand, vascular leakage with edema or hemorrhage could have been the result of respiratory distress and hypoxia (Busl and Greer 2010). Most streptococci are normal inhabitants of mammalian genitourinary and respiratory mucosae (Fulde and Valentin-Weigand 2012), although neither streptococcal species observed in our study have been previously isolated at this colony. Furthermore, adult animals of Guafo Island have not been screened for bacterial pathogens, and further studies are necessary to test if adult animals are a source of infection for pups.

The epidemic nature of the bronchopneumonia cases in SAFS pups is intriguing because previous health assessments have failed to record similar findings. Because it is likely that the pathogens associated with this outbreak have been present in this rookery without causing mortality, environmental factors may play an important role in this particular outbreak. For instance, the observed prevalence of pneumonia in Northern fur seal (Callorhinus ursinus) pups is higher in years with more precipitation, probably related to cold stress and disruption of pups' innate immunity (Spraker and Lander 2010). Some pinniped species may experience nutritional stress due to a reduction of pelagic fish availability during years of increased sea surface temperature (Banuet-Martinez et al. 2017). In California sea lions (Zalophus californianus), this stress appears to be related to lower levels of immunoglobulin A and immunoglobulin G (Banuet-Martinez et al. 2017). At the end of the 2016 breeding season at Guafo Island, there was an abrupt increase in sea surface temperature from 12 C to 15.2 C (National Aeronautics and Space Administration 2017). Acute nutritional stress and immunosuppression may have affected the health of SAFS adult females, facilitating the transfer of pathogenic bacteria and mites to their offspring.

We appreciate the logistical support of the Chilean Navy, artisanal fishermen of Quellon (Vessels Marimar II and Nautylus V), and the crews of the Chilean Navy Guafo Island lighthouse. We appreciate the field assistance of Rocio Carrasco and the support of María Carolina Silva-de la Fuente with parasite processing. This work was supported by The Rufford Small Grant Foundation, Morris Animal Foundation (Grant D16ZO-413), and the Society for Marine Mammalogy Small Grants in aid of research 2016. Work was conducted under permits of the Chilean Subsecretary of Fisheries Service (Res. Ex. 976, 2016) and the Institutional Animal Use and Care Committee of the University of Georgia (Project ID A2013 11-004-Y3-A0).

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