Stranding and mass mortality in humboldt penguins (Spheniscus humboldti), associated to HPAIV H5N1 outbreak in Chile.

The highly pathogenic Avian Influenza virus (HPAIV) H5N1 has caused a global outbreak affecting both wild and domestic animals, predominantly avian species. To date, cases of the HPAIV H5 Clade 2.3.4.4b in penguins have exclusively been reported in African Penguins. In Chile, the virus was confirmed in pelicans in December 2022 and subsequently spread across the country, affecting several species, including Humboldt penguins. This study aims to provide an overview of the incidents involving stranded and deceased Humboldt penguins and establish a connection between these events and HPAIV H5N1. Historical data about strandings between 2009 and 2023 was collected, and samples from suspected cases in 2023 were obtained to confirm the presence of HPAIV H5N1. Between January and August 2023, 2,788 cases of stranded and deceased penguins were recorded. Out of these, a total of 2,712 penguins deceased, evidencing a significative increase in mortality starting in early 2023 coinciding with the introduction and spreading of HPAIV H5N1 in the country. Thirty-seven events were categorized as mass mortality events, with the number of deceased penguins varying from 11 to 98. Most cases (97 %) were observed in the North of Chile. One hundred and eighty-one specimens were subjected to HPAIV diagnosis, four of which tested positive for HPAIV H5N1. Spatial analysis validates the correlation between mass mortality events and outbreaks of HPAIV in Chile. However, the limited rate of HPAIV H5N1 detection, which can be attributed to the type and quality of the samples, requiring further exploration.

Total infectome investigation of diphtheritic stomatitis in yellow-eyed penguins (Megadyptes antipodes) reveals a novel and abundant megrivirus.

First identified in 2002, diphtheritic stomatitis (DS) is a devastating disease affecting yellow-eyed penguins (Megadyptes antipodes, or hoiho in te reo Maori). The disease is associated with oral lesions in chicks and has caused significant morbidity and mortality. DS is widespread among yellow-eyed penguin chicks on mainland New Zealand yet appears to be absent from the subantarctic population. Corynebacterium spp. have previously been suspected as causative agents yet, due to inconsistent cultures and inconclusive pathogenicity, their role in DS is unclear. Herein, we used a metatranscriptomic approach to identify potential causative agents of DS by revealing the presence and abundance of all viruses, bacteria, fungi and protozoa - together, the infectome. Oral and cloacal swab samples were collected from presymptomatic, symptomatic and recovered chicks along with a control group of healthy adults. Two novel viruses from the Picornaviridae were identified, one of which - yellow-eyed penguin megrivirus - was highly abundant in chicks irrespective of health status but not detected in healthy adults. Tissue from biopsied oral lesions also tested positive for the novel megrivirus upon PCR. We found no overall clustering among bacteria, protozoa and fungi communities at the genus level across samples, although Paraclostridium bifermentans was significantly more abundant in oral microbiota of symptomatic chicks compared to other groups. The detection of a novel and highly abundant megrivirus has sparked a new line of inquiry to investigate its potential association with DS.

Four novel picornaviruses detected in Magellanic Penguins (Spheniscus magellanicus) in Chile.

Members of the Picornaviridae family comprise a significant burden on the poultry industry, causing diseases such as gastroenteritis and hepatitis. However, with the advent of metagenomics, a number of picornaviruses have now been revealed in apparently healthy wild birds. In this study, we identified four novel viruses belonging to the family Picornaviridae in healthy Magellanic penguins, a near threatened species. All samples were subsequently screened by RT-PCR for these new viruses, and approximately 20% of the penguins were infected with at least one of these viruses. The viruses were distantly related to members of the genera Hepatovirus, Tremovirus, Gruhelivirus and Crahelvirus. Further, they had more than 60% amino acid divergence from other picornaviruses, and therefore likely constitute novel genera. Our results demonstrate the vast undersampling of wild birds for viruses, and we expect the discovery of numerous avian viruses that are related to hepatoviruses and tremoviruses in the future.

Genomic Characterisation of a Novel Avipoxvirus Isolated from an Endangered Yellow-Eyed Penguin (Megadyptes antipodes).

Emerging viral diseases have become a significant concern due to their potential consequences for animal and environmental health. Over the past few decades, it has become clear that viruses emerging in wildlife may pose a major threat to vulnerable or endangered species. Diphtheritic stomatitis, likely to be caused by an avipoxvirus, has been recognised as a significant cause of mortality for the endangered yellow-eyed penguin (Megadyptes antipodes) in New Zealand. However, the avipoxvirus that infects yellow-eyed penguins has remained uncharacterised. Here, we report the complete genome of a novel avipoxvirus, penguinpox virus 2 (PEPV2), which was derived from a virus isolate obtained from a skin lesion of a yellow-eyed penguin. The PEPV2 genome is 349.8 kbp in length and contains 327 predicted genes; five of these genes were found to be unique, while a further two genes were absent compared to shearwaterpox virus 2 (SWPV2). In comparison with penguinpox virus (PEPV) isolated from an African penguin, there was a lack of conservation within the central region of the genome. Subsequent phylogenetic analyses of the PEPV2 genome positioned it within a distinct subclade comprising the recently isolated avipoxvirus genome sequences from shearwater, canary, and magpie bird species, and demonstrated a high degree of sequence similarity with SWPV2 (96.27%). This is the first reported genome sequence of PEPV2 from a yellow-eyed penguin and will help to track the evolution of avipoxvirus infections in this rare and endangered species.

Identification and Distribution of Novel Cressdnaviruses and Circular molecules in Four Penguin Species in South Georgia and the Antarctic Peninsula.

There is growing interest in uncovering the viral diversity present in wild animal species. The remote Antarctic region is home to a wealth of uncovered microbial diversity, some of which is associated with its megafauna, including penguin species, the dominant avian biota. Penguins interface with a number of other biota in their roles as marine mesopredators and several species overlap in their ranges and habitats. To characterize the circular single-stranded viruses related to those in the phylum Cressdnaviricota from these environmental sentinel species, cloacal swabs (n = 95) were obtained from King Penguins in South Georgia, and congeneric Adélie Penguins, Chinstrap Penguins, and Gentoo Penguins across the South Shetland Islands and Antarctic Peninsula. Using a combination of high-throughput sequencing, abutting primers-based PCR recovery of circular genomic elements, cloning, and Sanger sequencing, we detected 97 novel sequences comprising 40 ssDNA viral genomes and 57 viral-like circular molecules from 45 individual penguins. We present their detection patterns, with Chinstrap Penguins harboring the highest number of new sequences. The novel Antarctic viruses identified appear to be host-specific, while one circular molecule was shared between sympatric Chinstrap and Gentoo Penguins. We also report viral genotype sharing between three adult-chick pairs, one in each Pygoscelid species. Sequence similarity network approaches coupled with Maximum likelihood phylogenies of the clusters indicate the 40 novel viral genomes do not fall within any known viral families and likely fall within the recently established phylum Cressdnaviricota based on their replication-associated protein sequences. Similarly, 83 capsid protein sequences encoded by the viruses or viral-like circular molecules identified in this study do not cluster with any of those encoded by classified viral groups. Further research is warranted to expand knowledge of the Antarctic virome and would help elucidate the importance of viral-like molecules in vertebrate host evolution.

Identification of Circovirus Genome in a Chinstrap Penguin (Pygoscelis antarcticus) and Adélie Penguin (Pygoscelis adeliae) on the Antarctic Peninsula.

Circoviruses infect a variety of animal species and have small (~1.8-2.2 kb) circular single-stranded DNA genomes. Recently a penguin circovirus (PenCV) was identified associated with an Adélie Penguin (Pygoscelis adeliae) with feather disorder and in the cloacal swabs of three asymptomatic Adélie Penguins at Cape Crozier, Antarctica. A total of 75 cloacal swab samples obtained from adults and chicks of three species of penguin (genus: Pygoscelis) from seven Antarctic breeding colonies (South Shetland Islands and Western Antarctic Peninsula) in the 2015-2016 breeding season were screened for PenCV. We identified new variants of PenCV in one Adélie Penguin and one Chinstrap Penguin (Pygoscelis antarcticus) from Port Charcot, Booth Island, Western Antarctic Peninsula, a site home to all three species of Pygoscelid penguins. These two PenCV genomes (length of 1986 nucleotides) share > 99% genome-wide nucleotide identity with each other and share ~87% genome-wide nucleotide identity with the PenCV sequences described from Adélie Penguins at Cape Crozier ~4400 km away in East Antarctica. We did not find any evidence of recombination among PenCV sequences. This is the first report of PenCV in Chinstrap Penguins and the first detection outside of Ross Island, East Antarctica. Given the limited knowledge on Antarctic animal viral diversity, future samples from Antarctic wildlife should be screened for these and other viruses to determine the prevalence and potential impact of viral infections.

Herpesvirus Encephalitis in a Little Blue Penguin (Eudyptula minor).

An 11-day-old little blue penguin (Eudyptula minor) died unexpectedly. Prior to hatching, the egg experienced trauma and resultant defects were repaired. The chick hatched without complication and was clinically normal prior to death. Necropsy revealed congested lungs. Histologic examination showed moderate nonsuppurative encephalitis with focally extensive neuronal necrosis and intranuclear inclusions in neurons within necrotic foci. Herpesvirus DNA was detected in brain tissue with a generic herpesvirus polymerase chain reaction. Sanger sequencing demonstrated 100% and 98% sequence homology to sphenicid alphaherpesvirus 1 and penguin herpesvirus 2, respectively. In situ hybridization demonstrated large amounts of herpesvirus nucleic acid in intranuclear inclusions and neuronal nuclei. Combined histology, polymerase chain reaction, Sanger sequencing, and in situ hybridization results were most consistent with herpesviral encephalitis, most likely caused by sphenicid alphaherpesvirus 1. To our knowledge, this is the first report of a herpesvirus infection causing encephalitis in a penguin and the first report of herpesvirus in this species.

Avian Influenza H5N8 Outbreak in African Penguins (Spheniscus demersus), Namibia, 2019.

In January 2019, high mortalities were reported among African Penguins (Spheniscus demersus) in a breeding colony on Halifax Island, Namibia, Africa. Analysis of samples by reverse transcription quantitative PCR indicated the presence of highly pathogenic avian influenza (HPAI) subtype H5N8. Sequence analysis of the hemagglutinin and neuraminidase genes confirmed the presence of the virus in the birds and its high similarity to HPAI subtype H5N8 identified in South Africa in 2017. There have been no previous reports of HPAI H5N8 in Namibia.

Identification of a Novel Adélie Penguin Circovirus at Cape Crozier (Ross Island, Antarctica).

Understanding the causes of disease in Antarctic wildlife is crucial, as many of these species are already threatened by environmental changes brought about by climate change. In recent years, Antarctic penguins have been showing signs of an unknown pathology: a feather disorder characterised by missing feathers, resulting in exposed skin. During the 2018-2019 austral summer breeding season at Cape Crozier colony on Ross Island, Antarctica, we observed for the first time an Adélie penguin chick missing down over most of its body. A guano sample was collected from the nest of the featherless chick, and using high-throughput sequencing, we identified a novel circovirus. Using abutting primers, we amplified the full genome, which we cloned and Sanger-sequenced to determine the complete genome of the circovirus. The Adélie penguin guano-associated circovirus genome shares <67% genome-wide nucleotide identity with other circoviruses, representing a new species of circovirus; therefore, we named it penguin circovirus (PenCV). Using the same primer pair, we screened 25 previously collected cloacal swabs taken at Cape Crozier from known-age adult Adélie penguins during the 2014-2015 season, displaying no clinical signs of feather-loss disorder. Three of the 25 samples (12%) were positive for a PenCV, whose genome shared >99% pairwise identity with the one identified in 2018-2019. This is the first report of a circovirus associated with a penguin species. This circovirus could be an etiological agent of the feather-loss disorder in Antarctic penguins.

Penguin megrivirus, a novel picornavirus from an Adélie penguin (Pygoscelis adeliae).

The complete genome sequence of a novel megrivirus of the family Picornaviridae was determined from nucleic acid extracted from a pool of six faecal specimens of Adélie penguins. The samples were collected near Bellingshausen Station, King George Island of the South Shetland Islands, Antarctica. Penguin megrivirus is the first megrivirus with a predicted L protein. It has an L-3-5-4 genome layout, a type IV IRES, and a long 3' untranslated region of 668 nt.

Novel penguin Avian avulaviruses 17, 18 and 19 are widely distributed in the Antarctic Peninsula.

Three novel Avian avulavirus species were discovered and isolated during 2017 from Gentoo penguins (Pygoscelis papua) at Kopaitic island in the Northwestern region of the Antarctic Peninsula. The viruses were officially named as Avian avulavirus 17 (AAV17), Avian avulavirus 18 (AAV18) and Avian avulavirus 19 (AAV19), collectively referred to as penguin avulaviruses (PAVs). To determine whether these viruses are capable of infecting the three species of Pygoscelis spp. penguins (Gentoo, Adelie and Chinstrap) and assess its geographical distribution, serum samples were collected from seven locations across the Antarctic Peninsula and Southern Shetland Islands. The samples were tested by Hemagglutination inhibition assay using reference viruses for AAV17, AAV18 and AAV19. A total of 498 sera were tested, and 40 were positive for antibodies against AAV17, 20 for AAV18 and 45 for AAV19. Positive sera were obtained for the penguin's species for each virus; however, antibodies against AAV18 were not identified in Adelie penguins. Positive penguins were identified in all regions studied. Positive locations include Ardley Island and Cape Shirreff at Livingston Island (Southern Shetland Region); Anvers Island, Doumer Island and Paradise Bay in the Central Western region; and Avian Island at Southwestern region of the Antarctic Peninsula. The lowest occurrence was observed at the Southwestern region at Lagotellerie Island, where all samples were negative. On the other hand, Cape Shirreff and Paradise Bay showed the highest antibody titres. Field samples did not evidence cross-reactivity between viruses, and detection was significantly higher for AAV19 and lower for AAV18. This is the first serologic study on the prevalence of the novel Avian avulaviruses including different locations in the white continent. The results indicate that these novel viruses can infect the three Pygoscelis spp. penguins, which extend across large distances of the Antarctic Peninsula.

Antarctic Penguins as Reservoirs of Diversity for Avian Avulaviruses.

Wild birds harbor a huge diversity of avian avulaviruses (formerly avian paramyxoviruses). Antarctic penguin species have been screened for avian avulaviruses since the 1980s and, as such, are known hosts of these viruses. In this study, we screened three penguin species from the South Shetland Islands and the Antarctic Peninsula for avian avulaviruses. We show that Adelie penguins (Pygoscelis adeliae) are hosts for four different avian avulavirus species, the recently described avian avulaviruses 17 to 19 and avian avulavirus 10-like, never before isolated in Antarctica. A total of 24 viruses were isolated and sequenced; avian avulavirus 17 was the most common, and phylogenetic analysis demonstrated patterns of occurrence, with different genetic clusters corresponding to penguin age and location. Following infection in specific-pathogen-free (SPF) chickens, all four avian avulavirus species were shed from the oral cavity for up to 7 days postinfection. There was limited shedding from the cloaca in a proportion of infected chickens, and all but one bird seroconverted by day 21. No clinical signs were observed. Taken together, we propose that penguin species, including Antarctic penguins, may be the central reservoir for a diversity of avian avulavirus species and that these viruses have the potential to infect other avian hosts.IMPORTANCE Approximately 99% of all viruses are still to be described, and in our changing world, any one of these unknown viruses could potentially expand their host range and cause epidemic disease in wildlife, agricultural animals, or humans. Avian avulavirus 1 causes outbreaks in wild birds and poultry and is thus well described. However, for many avulavirus species, only a single specimen has been described, and their viral ecology and epidemiology are unknown. Through the detection of avian avulaviruses in penguins from Antarctica, we have been able to expand upon our understanding of three avian avulavirus species (avian avulaviruses 17 to 19) and report a potentially novel avulavirus species. Importantly, we show that penguins appear to play a key role in the epidemiology of avian avulaviruses, and we encourage additional sampling of this avian group.

Novel Avulaviruses in Penguins, Antarctica.

We identified 3 novel and distinct avulaviruses from Gentoo penguins sampled in Antarctica. We isolated these viruses and sequenced their complete genomes; serologic assays demonstrated that the viruses do not have cross-reactivity between them. Our findings suggest that these 3 new viruses represent members of 3 novel avulavirus species.

Genetically diverse herpesviruses in South American Atlantic coast seabirds.

Different herpesviruses have been associated with respiratory and enteric disease and mortality among seabirds and waterfowl. In 2011, a respiratory disease outbreak affected 58.3% (98/168) of the Magellanic penguins undergoing rehabilitation due to an oil spill off the southern Brazilian coast. Etiology was attributed to a novel herpesvirus identified by histopathology, immunohistochemistry, electron microscopy and molecular studies with partial DNA sequencing. Since migration, rehabilitation and translocation may facilitate the spread of pathogens between populations and trigger the onset of clinical disease in animals with latent infections, investigation of herpesvirus occurrence in asymptomatic seabirds was performed. Samples from free-ranging seabirds were collected in Argentinian Patagonia (Magellanic penguins) and the Abrolhos Archipelago in Brazil (Brown boobies, Masked boobies, Red-billed tropicbirds, White-tailed tropicbirds and South American tern). Furthermore, asymptomatic seabirds housed at the facility where the outbreak occurred were also sampled. In total, 354 samples from eight seabird species were analyzed by PCR for herpesvirus. Four different sequences of herpesviruses were identified, one in Yellow-nosed Albatross, one in Boobies and Tropicbirds and two in Magellanic penguins. Magellanic penguin herpesvirus 1 was identified during the penguin outbreak at the rehabilitation facility in Brazil, while Magellanic penguin herpesvirus 2 was recovered from free-ranging penguins at four reproduction sites in Argentina. Phylogenic analysis of the herpesviruses sequences tentatively identified suggested that the one found in Suliformes and the one associated with the outbreak are related to sequences of viruses that have previously caused seabird die-offs. These findings reinforce the necessity for seabird disease surveillance programs overall, and particularly highlight the importance of quarantine, good hygiene, stress management and pre-release health exams in seabirds undergoing rehabilitation.

DIPHTHERITIC STOMATITIS IN YELLOW-EYED PENGUINS (MEGADYPTES ANTIPODES) IN NEW ZEALAND.

Diphtheritic stomatitis is a seasonal disease that has been recognized as a syndrome in Yellow-eyed Penguin ( Megadyptes antipodes ) chicks in New Zealand for >10 yr. It was present in about 50% of 234 chicks examined since 2002 and is characterized by a thick serocellular exudate in the oral cavity of 1-4-wk-old chicks. The syndrome includes inanition, weight loss, and death in many affected birds. Microscopically, the lesions varied in severity. Most affected chicks had severe, locally extensive, ulcerative stomatitis with large amounts of exudate containing numerous bacteria; a smaller number had mild focal lesions with smaller amounts of exudate and bacteria. Although Corynebacterium amycolatum has been consistently isolated from the oral lesions, it was also present in the oral cavity of 34% of normal adult penguins and their chicks and is not known to possess diphtheritic toxins. A primary viral pathogen was therefore suspected, and intracytoplasmic inclusion bodies were occasionally seen in oral mucosal epithelial cells. No herpesvirus DNA was detected with PCR. Avipoxvirus DNA and an unidentified virus-like agent were detected in some early oral lesions, but could not be confirmed in subsequent testing. Electron microscopy on early affected epithelium with intracytoplasmic inclusion bodies was unrewarding. Our findings raise the possibility that the disease is caused by an unknown primary virus infection followed by secondary Corynebacterium invasion, but this requires confirmation. The means of transmission has not been established but insect vectors are suspected.

A novel alphaherpesvirus associated with fatal diseases in banded Penguins.

A novel avian alphaherpesvirus, preliminarily designated sphenicid alphaherpesvirus 1 (SpAHV-1), has been independently isolated from juvenile Humboldt and African penguins (Spheniscus humboldti and Spheniscus demersus) kept in German zoos suffering from diphtheroid oropharyngitis/laryngotracheitis and necrotizing enteritis (collectively designated as penguin-diphtheria-like disease). High-throughput sequencing was used to determine the complete genome sequences of the first two SpAHV-1 isolates. SpAHV-1 comprises a class D genome with a length of about 164 kbp, a G+C content of 45.6 mol% and encodes 86 predicted ORFs. Taxonomic association of SpAHV-1 to the genus Mardivirus was supported by gene content clustering and phylogenetic analysis of herpesvirus core genes. The presented results imply that SpAHV-1 could be the primary causative agent of penguin-diphtheria-like fatal diseases in banded penguins. These results may serve as a basis for the development of diagnostic tools in order to investigate similar cases of penguin diphtheria in wild and captive penguins.

Evidence for the Introduction, Reassortment, and Persistence of Diverse Influenza A Viruses in Antarctica.

Avian influenza virus (AIV) surveillance in Antarctica during 2013 revealed the prevalence of evolutionarily distinct influenza viruses of the H11N2 subtype in Adélie penguins. Here we present results from the continued surveillance of AIV on the Antarctic Peninsula during 2014 and 2015. In addition to the continued detection of H11 subtype viruses in a snowy sheathbill during 2014, we isolated a novel H5N5 subtype virus from a chinstrap penguin during 2015. Gene sequencing and phylogenetic analysis revealed that the H11 virus detected in 2014 had a >99.1% nucleotide similarity to the H11N2 viruses isolated in 2013, suggesting the continued prevalence of this virus in Antarctica over multiple years. However, phylogenetic analysis of the H5N5 virus showed that the genome segments were recently introduced to the continent, except for the NP gene, which was similar to that in the endemic H11N2 viruses. Our analysis indicates geographically diverse origins for the H5N5 virus genes, with the majority of its genome segments derived from North American lineage viruses but the neuraminidase gene derived from a Eurasian lineage virus. In summary, we show the persistence of AIV lineages in Antarctica over multiple years, the recent introduction of gene segments from diverse regions, and reassortment between different AIV lineages in Antarctica, which together significantly increase our understanding of AIV ecology in this fragile and pristine environment. IMPORTANCE: Analysis of avian influenza viruses (AIVs) detected in Antarctica reveals both the relatively recent introduction of an H5N5 AIV, predominantly of North American-like origin, and the persistence of an evolutionarily divergent H11 AIV. These data demonstrate that the flow of viruses from North America may be more common than initially thought and that, once introduced, these AIVs have the potential to be maintained within Antarctica. The future introduction of AIVs from North America into the Antarctic Peninsula is of particular concern given that highly pathogenic H5Nx viruses have recently been circulating among wild birds in parts of Canada and the Unites States following the movement of these viruses from Eurasia via migratory birds. The introduction of a highly pathogenic influenza virus in penguin colonies within Antarctica might have devastating consequences.

Genetic and Molecular Epidemiological Characterization of a Novel Adenovirus in Antarctic Penguins Collected between 2008 and 2013.

Antarctica is considered a relatively uncontaminated region with regard to the infectious diseases because of its extreme environment, and isolated geography. For the genetic characterization and molecular epidemiology of the newly found penguin adenovirus in Antarctica, entire genome sequencing and annual survey of penguin adenovirus were conducted. The entire genome sequences of penguin adenoviruses were completed for two Chinstrap penguins (Pygoscelis antarctica) and two Gentoo penguins (Pygoscelis papua). The whole genome lengths and G+C content of penguin adenoviruses were found to be 24,630-24,662 bp and 35.5-35.6%, respectively. Notably, the presence of putative sialidase gene was not identified in penguin adenoviruses by Rapid Amplification of cDNA Ends (RACE-PCR) as well as consensus specific PCR. The penguin adenoviruses were demonstrated to be a new species within the genus Siadenovirus, with a distance of 29.9-39.3% (amino acid, 32.1-47.9%) in DNA polymerase gene, and showed the closest relationship with turkey adenovirus 3 (TAdV-3) in phylogenetic analysis. During the 2008-2013 study period, the penguin adenoviruses were annually detected in 22 of 78 penguins (28.2%), and the molecular epidemiological study of the penguin adenovirus indicates a predominant infection in Chinstrap penguin population (12/30, 40%). Interestingly, the genome of penguin adenovirus could be detected in several internal samples, except the lymph node and brain. In conclusion, an analysis of the entire adenoviral genomes from Antarctic penguins was conducted, and the penguin adenoviruses, containing unique genetic character, were identified as a new species within the genus Siadenovirus. Moreover, it was annually detected in Antarctic penguins, suggesting its circulation within the penguin population.

Herpesvirus-like respiratory infection in African penguins Spheniscus demersus admitted to a rehabilitation centre.

Rehabilitation is an important strategy for the conservation of the Endangered African penguin Spheniscus demersus, and disease has been raised as a concern in the management of the species, both in the wild and in rehabilitation centres. We report 8 cases of herpesvirus-like respiratory infection in African penguin chicks undergoing rehabilitation between 2010 and 2013 at a facility in Cape Town, South Africa. Infection was confirmed through the identification of viral inclusions in the tracheal epithelium and demonstration of particles consistent with herpesvirus by electron microscopy, whereas virus isolation in eggs, serology and PCR testing failed to detect the virus. Only penguin chicks were affected; they were in poor body condition, and in 2 cases infection occurred prior to admission to the rehabilitation centre. The role played by the herpesvirus-like infection in the overall respiratory disease syndrome is uncertain, due to identification of lesions in only a small proportion of the chicks as well as to the occurrence of other concurrent pathological processes. Further studies are advised to characterise the specific virus involved through the development of sensitive diagnostic methods and to clarify the epidemiology and significance of these infections in wild African penguins.