Human Parainfluenza Virus Type 3 in Wild Nonhuman Primates, Zambia

Human parainfluenza virus type 3 (HPIV3) genome was detected in 4 baboons in Zambia. Antibody for HPIV3 was detected in 13 baboons and 6 vervet monkeys in 2 distinct areas in Zambia. Our findings suggest that wild nonhuman primates are susceptible to HPIV3 infection.

Human parainfluenza virus type 3 (HPIV3) genome was detected in 4 baboons in Zambia. Antibody for HPIV3 was detected in 13 baboons and 6 vervet monkeys in 2 distinct areas in Zambia. Our findings suggest that wild nonhuman primates are susceptible to HPIV3 infection.
H uman parainfluenza viruses (HPIVs) (family Paramyxoviridae) are major causes of lower respiratory tract infections in infants and elderly persons. HPIVs are second to respiratory syncytial virus as the cause of hospitalizations for lower respiratory tract infections (1,2) and account for 6.8% of all hospitalizations for fever or acute respiratory illness in children <5 years of age (3). Among the 4 serotypes, HPIV3 (genus Respirovirus) causes particularly severe disease, including bronchiolitis and pneumonia (1)(2)(3). In addition to young children, HPIV3 poses a threat to the elderly and to immunocompromised adults. HPIV3 infection also causes severe illness leading to death (35%-75% death rate) in patients receiving hematopoietic stem cell transplants (4,5). Although the virus is distributed worldwide and maintained in human communities, its epidemiology in Africa is unclear.
Nonhuman primates, the closest living relatives of humans, are susceptible to paramyxoviruses that cause respiratory disease in humans. Recently, other researchers reported infections with human respiratory syncytial virus and human metapneumovirus in wild nonhuman primates in Africa (6,7). Therefore, as a first step in determining the pervasiveness of infection in African wild nonhuman primates, we screened these animals for paramyxovirus in Zambia. The HPIV3 genome was identified by seminested broad-spectrum reverse transcription PCR (RT-PCR). Thereafter, we investigated HPIV3 infection in wild nonhuman primates by using molecular and serologic methods.

The Study
Baboons and vervet monkeys live side by side with humans in game management areas in Zambia, and this situation often leads to high levels of human-baboon/monkey conflicts. The Zambia Wildlife Authority is mandated by the Zambian government to control the large numbers of these animals. We collected tissues and serum samples from  Table. The species were identified on the basis of morphologic characters and the mitochondrial cytochrome b (cytb) gene sequence. The complete cytb gene was amplified from spleen DNA of Papio spp. baboons with the primer set papio cytb1F (5′-GATACGAAAAACCATC-GCTGT-3′) and papio cytb2R (5′-GCTCCATTTCTG-GTTTACAAG-3′), as described (8), and from spleen DNA of Chlorocebus spp. monkeys with the primer set chlorocebus cytb1F (5′-TGATATGAAAAACCACCGTTGT-3′) and chlorocebus cytb2R (5′-GCTTTCTTTCTGAGTT-GTCCTAGG-3′), designed in this study.
Total RNA was extracted from 189 spleen tissue samples by using TRIzol reagent (Life Technologies, Carlsbad, CA, USA) and screened for paramyxoviruses by seminested broad-spectrum RT-PCR of paramyxovirus polymerase (L) genes (9). Amplification was carried out with the primers PAR-F1, PAR-F2, and PAR-R. Seminested RT-PCR was positive in only 1 chacma baboon sample. The PCR product (584 bp) was subjected to direct sequence analysis, and the identified paramyxovirus was tentatively named ZMLS/2011. BLAST analysis (http://blast.ncbi.nlm.nih. gov/blast.cgi) indicated that ZMLS/2011 shared 98% nt identity with the HPIV3 L gene. To increase the sensitivity of HPIV3 genome detection, we screened all 189 RNA samples by RT-PCR, using the HPIV3 L gene-specific primer sets HPIV3 L1F (5′-ATGGGAGAATTCTTCCT-CAAGCTC-3′) and HPIV3 L2R (5′-AATGCRGCAACT-GATGGATCACC-3′). An HPIV3 genome was detected in 3 (6%) of 50 chacma baboon samples and in 1 (2%) of 50 yellow baboon samples but not from any of the 89 vervet monkey samples. Nucleotide sequences of all 4 amplicons (367 bp) were identical to ZMLS/2011. In an attempt to isolate virus from RT-PCR-positive spleen, Vero cells cultured in minimum essential medium supplemented with trypsin were injected with tissue homogenates; however, after 3 passages, cytopathic effects were not observed. Viral RNA was also not detected in the culture supernatants from the cells.
To   animals in the Mfuwe and Livingstone regions. All 4 baboons positive for the HPIV3 genome were negative for HPIV3 antibodies (data not shown), suggesting that, at the time the samples were taken, these HPIV3 antibody-negative baboons might have been in the acute stage of infection, before a detectable immune response had developed.

Conclusions
We identified wild baboons positive for HPIV3 by using molecular and serologic analyses. Seropositive vervet monkeys also were found. These nonhuman primates are widely distributed in Africa (8), and their habitats overlap those of humans, mainly in rural areas. A survey performed in Kenya of humans with influenza-like illness and severe acute respiratory illness showed that 9.4% were positive for HPIVs (12). However, HPIV3 infection in humans in Africa has been poorly studied, making ZMLS/2011 difficult to compare with an epidemic strain of HPIV3 among humans. The nonhuman primates sampled in this study live side by side with humans. In addition, many tourists visit the Livingstone region and, in some instances, are harmed by the nonhuman primates attempting to grab food carried by humans. Detection of HPIV3 in wild nonhuman primates might be related to these contacts. Further epidemiologic studies of humans and wild nonhuman primates are needed to determine whether HPIV3 is transmitted between humans and wild nonhuman primates.
Serologic evidence of HPIV3 infection was obtained from baboons and vervet monkeys in 2 distinct geographic areas of Zambia, but little is known about HPIV3 infection in wild nonhuman primates. In 1963, simian agent 10 (also known as simian virus 10) was isolated from the mouth of a Samango monkey (Cercopithecus mitis) in a laboratory in South Africa (13). Complete genome sequence analysis showed simian agent 10 as a strain of HPIV3 (14). Experimental infections showed that many nonhuman primatesincluding chimpanzees; macaques; and squirrel, owl, patas, and rhesus monkeys-are sensitive to HPIV3 infection (1,15). These previous reports support our finding that wild nonhuman primates are susceptible to HPIV3 infection.