A genetic comparison of two alleged subspecies of Philippine cynomolgus macaques

Abstract

Two subspecies of cynomolgus macaques (Macaca fascicularis) are alleged to co-exist in the Philippines, M. f. philippensis in the north and M. f. fascicularis in the south. However, genetic differences between the cynomolgus macaques in the two regions have never been studied to document the propriety of their subspecies status. We genotyped samples of cynomolgus macaques from Batangas in southwestern Luzon and Zamboanga in southwestern Mindanao for 15 short tandem repeat (STR) loci and sequenced an 835 bp fragment of the mtDNA of these animals. The STR genotypes were compared with those of cynomolgus macaques from southern Sumatra, Singapore, Mauritius and Cambodia, and the mtDNA sequences of both Philippine populations were compared with those of cynomolgus macaques from southern Sumatra, Indonesia and Sarawak, Malaysia. We conducted STRUCTURE and PCA analyses based on the STRs and constructed a median joining network based on the mtDNA sequences. The Philippine population from Batangas exhibited much less genetic diversity and greater genetic divergence from all other populations, including the Philippine population from Zamboanga. Sequences from both Batangas and Zamboanga were most closely related to two different mtDNA haplotypes from Sarawak from which they are apparently derived. Those from Zamboanga were more recently derived than those from Batangas, consistent with their later arrival in the Philippines. However, clustering analyses do not support a sufficient genetic distinction of cynomolgus macaques from Batangas from other regional populations assigned to subspecies M. f. fascicularis to warrant the subspecies distinction M. f. philippensis.

Keywords: Macaca fascicularis; mitochondrial DNA; short tandem repeat (STR); subspecies.

Figure 1

Map of the distribution of M. fascicularis illustrating locations of three major and seven minor (on offshore islands) subspecies based on Groves (2001). Names in capital letters with dark arrows indicate the smallest geographical unit (e.g., country, province, island, etc.) within which samples used in this analysis originated. Arrows indicate routes of dispersal of M. f. philippensis, first, across the Palawan Islands and M. f. fascicularis, much later, across the Sulu Archipelago to the Philippines.

Figure 2

Principal Coordinates Analysis (PCA) based on genotypes at 15 STR loci in seven regional populations of cynomolgus macaques. Ellipses indicate the 95% confidence limits for each regional population and grid intervals are in units 0f 0.5.

Figure 3

A. STRUCTURE plot of three Philippine populations (Batangas, Corregidor and Zamboanga) with K=2 based on genotypes at 15 STR loci. Batangas and Zamboanga are well differentiated in the plot with Corregidor showing admixture of approximately 25% and 75% with each of the two populations. B. STRUCTURE plot of all seven populations of cynomolgus macaques based on STRs when K=3. Note that the three Philippine populations for a single group that is well differentiated from populations from Sumatra and Mauritius.

Figure 4

Maximum Likelihood tree for a 697 bp fragment of mtDNA of five regional populations of cynomolgus macaques based on the Tamura-Nei model with 500 replicates. Branches corresponding to partitions reproduced in fewer than 50% bootstrap replicates were collapsed. Bootstrap values are provided for each branch. Sequences with “Fas” prefixes are reference sequences representing previously identified mtDNA haplogroups in cynomolgus macaques (Smith et al., 2007). Each such reference is followed by the identification number of the sample representative of that haplogoup whose prefix identifies the country of origin of that sample. Sequences with prefixes “Cor”, “Luz”, “Zam”, “Sar”, “Sum”, “Phi”, “Viet”, “Mcy”, “Maur”, and “Imcy” are sequences of samples from Corregidor, Luzon (Batangas), Zamboanga, Sarawak, Sumatra, Philippines (Mindanao), Vietnam, Malaysia, Mauritius and Indonesia, respectively. The tree is rooted with a sequence from M. sylvanus.

Figure 5

Median-Joining haplotype network based on mtDNA sequences employed in Figure 5, above, and rooted with a sequence from M. sylvanus. The reference sequences are identified by both their sample numbers and haplogroup acronym of each reference sample. Note: The sequence of sample number 10 (**) from Zamboanga is identical to the haplogroup reference sequence Fas2a2 and is shared with Corregidor and the sequence of sample number 50 from Batangas is shared with Corregidor.